Lab 15: Population Genetics - eScience Labs

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Lab 15: Population Genetics - eScience Labs

Lab Manual

Introductory Biology (Version 1.4)

© 2011 eScience Labs, LLC

All rights reserved

www.esciencelabs.com • 888.375.5487


Table of Contents:

Introduction:

Lab 1: The Scientific Method

Lab 2: Writing a Lab Report

Lab 3: Data Measurement

Lab 4: Introduction to the Microscope

Biological Processes:

Lab 5: The Chemistry of Life

Lab 6: Diffusion

Lab 7: Osmosis

Lab 8: Respiration

Lab 9: Enzymes

The Cell:

Lab 10: Cell Structure & Function

Lab 11: Mitosis

Lab 12: Meiosis

Lab 13: DNA & RNA

Lab 14: Mendelian Genetics

Lab 15: Population Genetics

3


Common Labware found in ESL Kits

4


Lab Safety

Always follow the instructions in your laboratory manual and these general rules:

eScience Labs, Inc. designs every kit with safety as our top priority.

Nonetheless, these are science kits and contain items which must be

handled with care. Safety in the laboratory always comes first!

Lab preparation



Please thoroughly read the lab exercise before starting!

If you have any doubt as to what you are supposed to be doing and how to do it safely,

please STOP and then:

Double‐check the manual instructions.

Check www.esciencelabs.com for updates and tips.

Contact us for technical support by phone at 1‐888‐ESL‐Kits (1‐888‐375‐5487) or by email

at Help@esciencelabs.com.


Read and understand all labels on chemicals.

If you have any questions or concerns, refer to the Material Safely Data Sheets (MSDS)

available at www.esciencelabs.com. The MSDS lists the dangers, storage requirements,

exposure treatment and disposal instructions for each chemical.


Consult your physician if you are pregnant, allergic to chemicals, or have other medical

conditions that may require additional protective measures.

Proper lab attire





Remove all loose clothing (jackets, sweatshirts, etc.) and always wear closed‐toe shoes.

Long hair should be pulled back and secured and all jewelry (rings, watches, necklaces,

earrings, bracelets, etc.), should be removed.

Safety glasses or goggles should be worn at all times. In addition, wearing soft contact

lenses while conducting experiments is discouraged, as they can absorb potentially

harmful chemicals.

When handling chemicals, always wear the protective goggles, gloves, and apron

provided.

5


Performing the experiment




Do not eat, drink, chew gum, apply cosmetics or smoke while conducting an experiment.

Work in a well ventilated area and monitor experiments at all times, unless instructed

otherwise.

When working with chemicals:

Never return unused chemicals to their original container or place chemicals in an

unmarked container.

Always put lids back onto chemicals immediately after use.

Never ingest chemicals. If this occurs, seek immediate help.

Call 911 or “Poison Control” 1‐800‐222‐1222



Never pipette anything by mouth.

Never leave a heat source unattended.

If there is a fire, evacuate the room immediately and dial 911.

Lab Clean‐up and Disposal






If a spill occurs, consult the MSDS to determine how to clean it up.

Never pick up broken glassware with your hands. Use a broom and a dustpan and discard

in a safe area.

Do not use any part of the lab kit as a container for food.

Safely dispose of chemicals. If there are any special requirements for disposal, it will

be noted in the lab manual.

When finished, wash hands and lab equipment thoroughly with soap and water.

Above all, USE COMMON SENSE!

6


Approximate Time and Additional Materials Needed for Each Lab

** Note: If you are allergic to latex, please contact us and we will send you vinyl gloves**

Introduction:

Lab 1: The Scientific Method

Time: 1 hour

Materials: None

Lab 2: Writing a Lab Report

Time: 1 hour (plus 24 hours preparation time and 7‐10 days for observation)

Materials: Paper towels, water, masking tape

Lab 3: Data Measurement

Time: 1 hour

Materials: Water

Lab 4: Introduction to the Microscope

Time: 1 hour

Materials: Access to ESL’s Student Portal

Biological Processes:

Lab 5: The Chemistry of Life

Time: 1 hour (plus 24 hours preparation time)

Materials: Variety of household substances, plastic wrap, water, cutting

utensil

Lab 6: Diffusion

Time: 1.5 hours

Materials: Water, watch or timer , viscous liquid from cupboard

Lab 7: Osmosis

Time: 1 hour (plus 3 hours for observation)

Materials: Water, watch or timer, several types of potatoes, cutting utensil,

paper towel

Lab 8: Respiration

Time: 1 hour (plus 2 hours preparation time)

Materials: Water, watch or timer, paper towel

Lab 9: Enzymes

Time: 1 hour (plus 2 hours preparation time)

Materials: Water, watch or timer, string, ice, hot water, paper towel,

ginger root, at least 2 other food sources (potato, apple, etc.)

The Cell:

Lab 10: Cell Structure & Function

Time: 1 hour (plus 24 hours for observation)

Materials: Water, square plastic food storage container, mixing bowl, house

hold items for use as cell structures (plums, raisins, etc.)

Lab 11: Mitosis

Time: 1 hour

Materials: None

7


Lab 12: Meiosis

Time: 1.5 hours

Materials: Blue and red markers

Lab 13: DNA & RNA

Time: 2 hours

Materials: Fruit, scissors

Lab 14: Mendelian Genetics

Time: 1.5 hours

Materials: None

Lab 15: Population Genetics

Time: 1.5 hours

Materials: None

8


Additional Online Content Found at www.esciencelabs.com

Introduction:

ESL Safety Video

ESL Scientific Processes Video

How Big Is It?

Introduction to the Microscope

Measuring Volume Using a

Graduated Cylinder

Unit Conversions

Biological Processes:

The Cell:

ESL Biological Processes Video

The Structure of an Atom

Acid/Base Reactions

Diffusion and Osmosis Tutorial

Docking Tutorial

Log on to the Student Portal using

these easy steps:

Visit our website,

www.esciencelabs.com, and click on

the green button (says “Register or

Login”) on the top right side of the

page. From here, you will be taken to

a login page. If you are registering

your kit code for the first time, click

the “create and account” hyperlink.

Locate the kitcode, located on a label

on the inside of the kit box lid. Enter

this, along with other requested information

into the online form to create

your user account. Be sure to keep

track of your username and password

as this is how you will enter the Student

Portal for future visits. This establishes

your account with the

eScience Labs’ Student Portal.

Have fun!

ESL Cell Video

Cell Structure Crossword Puzzle

Interactive Videos of Meiosis

Interactive Videos of Mitosis

Nature’s Review of RNA

DNA Transcription & Translation

9


The Cell (continued):

How Mutations Work

Riken Center’s Developmental Biology Stem Cell Videos

A Typical Animal Cell

Construction of the Cell Membrane

The Cell Cycle

Cell Division

DNA Extraction Virtual Lab

Additional Resources:

Stop Watch

Conversion Tables

10


Introduction

Lab 1

The Scientific Method

11


Lab 1 : Scientific Method

Concepts to explore:

Concepts to explore:


Testable observations


Data collection


Hypothesis


Analysis


Null hypothesis


Experimental approach


Variables


Controls

Introduction

What is science? You have likely taken several classes throughout your career as a student, and know

that it is more than just chapters in a book. Science is a process that uses evidence to understand the

history of the natural world and how it works. It is constantly changing as we understand more about

the natural world, and continues to advance the understanding of the universe. Science begins with

observations that can be measured in some way so that data can be collected in a useful manner by following

the scientific method.

Have you ever wondered why the sky is blue or why a plant grows toward a window? If so, you have already

taken the first step down the road of discovery. No matter what the question, the scientific

method can help find an answer (or more than one answer!). Following the scientific method helps to

insure scientists can minimize bias when testing a theory. It will help you to collect and organize information

in a useful way, looking for connections and patterns in the data. As an experimenter, you

should use the scientific method as you conduct the experiments throughout this manual.

Figure 1: The process of the scientific method

13


Lab 1 : Scientific Method

The scientific method process begins with the formulation of a

hypothesis – a statement of what the experimenter thinks will

happen in certain situations. A hypothesis is an educated guess –

a proposed explanation for an event based on observation(s). A

null hypothesis is a testable statement, that if proven true means

the hypothesis was incorrect. Both statements must be testable,

but only one can be true. Hypotheses are typically written in an

if/then format, such as:

Hypothesis:

If nutrients are added to soil, then plants grown in it will

grow faster than plants without added nutrients in the soil.

Figure 2: What affects plant growth?

Null hypothesis:

If nutrients are added to the soil, then the

plants will grow the same as plants in soil

without added nutrients.

There are often many ways to test a hypothesis.

When designing an experiment to test a hypothesis

there are three rules to follow:

If plants grow quicker when nutrients are added,

then the hypothesis is accepted and the null

hypothesis is rejected.

1. The experiment must be replicable.

2. Only test one variable at a time.

3. Always include a control.

Variables are defined and measurable components of an experiment. Controlling the variables in an

experiment allows the scientist to quantitate the changes that occur so that results can be measured

and conclusions drawn. There are three types of variables:

Independent Variable: The variable that the scientist changes to a predetermined value

in order to test the hypothesis. There can only be one independent variable in each

experiment in order to pinpoint the change that affects the outcome of the experiment.

Dependent Variable: This variable is measured in regards to conditions of the independent

variable—it depends on the independent variable. There can be more than

one dependent variable in each experiment.

14


Lab 1 : Scientific Method

Controlled Variable: This variable, or variables (there could be many) reflect the factors

that could influence the results of the experiment, but are not the planned changes the

scientist is expecting (by changing the independent variable). These variables must be

controlled so that the results can be associated with some change in the independent

variable.

When designing the experiment, establish a clear and concise procedure. Controls must be identified to

eliminate compounding changes that can influence the results. Often times, the hardest part of designing

an experiment is not figuring out how to test the one factor you focus on, but in trying to eliminate

the often hidden influences that can skew results. Taking notes when conducting an experiment is important,

whether it is recording the temperature, humidity, time of day, or another environmental condition

that may have an impact on the results. Also remember that replication is fundamental to scientific

experiments. Before drawing conclusions, make sure your data is repeatable. In other words, make

sure the experiment provides significant results over multiple trials.

Often, the best way to organize data for analysis is as a table or a graph. Remember, any table or graph

should be able to stand on its own. In other words, another scientist should be able to pick up the table

or graph and have all of the information necessary to interpret it, with no other information.

Table: A well‐organized summary of data collected. Only include information relevant to the hypothesis

(e.g. don’t include the color of the plant because it’s not relevant to what is being tested). Always

include a clearly stated title, label your columns and rows and include the units of measurement.

For our example:

Table 1: Plant Growth with and without Added Nutrients

Variable Height Wk1 (mm) Height Wk 2 (mm) Height Wk 3 (mm) Height Wk 4 (mm)

Control

(without nutrients)

Independent

(with nutrients)

3.4 3.6 3.7

3.5 3.7 4.1 4.6

4.0

Graph: A visual representation of the relationship between the independent and dependent variable.

Graphs are useful in identifying trends and illustrating findings. Rules to remember:



The independent variable is always graphed on the x‐axis (horizontal), with the dependent

variable on the y axis (vertical).

Use appropriate numerical spacing when plotting the graph, with the lower numbers

starting on both the lower and left hand corners.

Always use uniform or logarithmic intervals. For example, if you begin by numbering, 0,

10, 20, do not jump to 25 then to 32.

15


Lab 1 : Scientific Method



Title the graph and both the x and y axes such that they correspond to the table from

which they come. For example, if you titled your table “Heart rate of those who eat

vegetables and those who do not eat vegetables”, be sure to title the graph the same.

Determine the most appropriate type of graph. Typically, line and bar graphs are the

most common.

Line graph: Shows the relationship between variables using plotted points that are connected with a

line. There must be a direct relationship and dependence between each point connected.

More than one set of data can be presented on a line graph. Figure 3 uses the data from our

previous table:

Height

(mm)

Figure 3: Plant growth, with and without nutrients, over time

Bar graph: Used to compare results that are independent from each other, as opposed to a continuous

series. Since the results from our previous example are continuous, they are not appropriate for a bar

graph.

Figure 4 shows the top speeds of four cars. Since there is no relationship between each car, each result

is independent and a bar graph is appropriate.

16


Lab 1 : Scientific Method

Speed (kph)

Figure 4: Top speed for Cars A, B, C, and D

Interpretation: Based on the data you collected, is your hypothesis supported or refuted? Based on the

data, is the null hypothesis supported or refuted? If the hypothesis is supported, are there other variables

which should be examined? For instance, was the amount of water and sunlight consistent between

groups of plants ‐ or, were all four cars driven on the same road?

Exercise 1:

Dissolved oxygen is oxygen that is trapped in a fluid, such as water. Since virtually every living organism

requires oxygen to survive, it is a necessary component of water systems such as streams, lakes and rivers

in order to support aquatic life. The dissolved oxygen is measure in units of ppm—or parts per million.

Examine the data in Table 2 showing the amount of dissolved oxygen present and the number of

fish observed in the body of water the sample was taken from; finally, answer the questions below.

17


Lab 1 : Scientific Method

Table 2: Water quality vs. fish population

Dissolved Oxygen (ppm) 0

2

4

6

8

10

12

14

16

18

Number of Fish Observed 0

1

3

10

12

13

15

10

12

13

1. Based on the information in Table 2, what patterns do you observe?

2. Develop a hypothesis relating to the amount of dissolved oxygen measured in the water

sample and the number of fish observed in the body of water.

3. What would your experimental approach be to test this hypothesis?

4. What are the independent and dependent variables?

5. What would be your control?

6. What type of graph would be appropriate for this data set? Why?

7. Graph the data from the table above.

18


Lab 1 : Scientific Method

8. Interpret the data from the graph made in Question 7.

Exercise 2:

Determine which of the following observations are testable. For those that are testable:

Write a hypothesis and null hypothesis

What would be your experimental approach?

What are the dependent and independent variables?

What is your control?

How will you collect your data?

How will you present your data (charts, graphs, types)?

How will you analyze your data?

1. When a plant is placed on a window sill, it grows faster than when it is placed on a coffee

table in the middle of the living room.

2. The teller at the bank with brown hair and brown eyes and is taller than the other tellers.

3. I caught four fish at the seven o’clock in the morning but didn’t catch any at noon.

4. The salaries at Smith and Company are based on the number of sales and Billy makes 3,000

dollars more than Joe.

19


Lab 1 : Scientific Method

5. When Sally eats healthy foods and exercises regularly, her blood pressure is lower than

when she does not exercise and eats fatty foods.

6. The Italian restaurant across the street closes at 9 pm but the one two blocks away closes at

10 pm.

7. Bob bought a new blue shirt with a golf club on the back for twenty dollars.

8. For the past two days the clouds have come out at 3 pm and it has started raining at 3:15

pm.

9. George did not sleep at all last night because he was up finishing his paper.

10. Ice cream melts faster on a warm summer day than on a cold winter day.

11. How can you apply scientific method to an everyday problem? Give one example.

20


Introduction

Lab 2

Writing a Lab Report

21


Lab 2: Writing a Lab Report

Concepts to explore:




What is a lab report?

The parts of a lab report

How to write a lab report

Introduction

A lab report is a scientific paper describing an experiment, how it was

done and the results of the study.

Experiments are performed to test whether what one thinks may happen,

actually does. The lab report lays out the results of the experiment

and can be used to communicate the findings to other scientists.

It allows the findings of one scientist to be examined, replicated,

refuted or supported by another scientist. Though most lab

reports go unpublished, it is important to write a report that accurately

characterizes the experiment performed.

Even if what is described never reaches the public or the scientific

community, the report lays the foundation for other experiments. It

also provides a written record of what was done, so that others can understand

what the investigator was thinking and doing.

Figure 1: Lab reports are an

essential part of science,

providing a means of reporting

experimental findings

Parts of a Lab Report:

Title: A short statement summarizing the topic of the report.

Abstract: A brief summary of the methods, results and conclusions. It should not exceed 200 words

and should be the last part written.

Introduction: This is an overview of why the experiment was conducted. There are three key parts:

Background: Provides an overview of what is already known and what questions remain unresolved

regarding the topic of the experiment. Assume the reader needs a basic introduction to

23


Lab 2: Writing a Lab Report

the topic and provide the information necessary for them to understand

why and how the experiment was performed.

Objective: Explain the purpose of the experiment. For example; “I

want to determine if taking baby aspirin every day prevents second

heart attacks”.

Hypothesis: This is your “guess” as to what will happen when you

do the experiment.

Materials and Methods: These are detailed descriptions of what was used

Figure 2: Follow the guidelines

in this introduction

to conduct the experiment, what was actually done (step by step) and how

it was done. The description should be exact enough that someone reading

the report can replicate the experiment. Make sure to include all the

when writing a lab report.

equipment and supplies used, even they seem obvious and did not seem to play a large role. When

describing the methods, go in order from the first step to the last. Do not list the procedures used in a

numerical fashion, but write them in complete sentences and paragraphs, much like you would if

speaking.

Results: This is the data obtained from the experiment. This section should be clear, concise and to

the point. In this section tables and graphs are often appropriate and frequently are the best way to

present the data. Do not include any interpretations, only the raw data.

Discussion: This is where the scientist (you) can interpret the data you obtained and draw conclusions.

Was your hypothesis (“guess”) supported or refuted? Discuss what these findings mean, look at common

themes, relationships and points that perhaps generate more questions. If fewer second heart

attacks were reported when baby aspirin was taken, but only in women, this would lead to additional

questions. When appropriate, discuss outside factors (i.e. temperature, time of day, etc.) that may

have played a role in the experiment and what could be done to control those in future experiments.

Conclusion: A short, pointed summary that states what has been learned from this experiment.

References: Any articles, books, magazines, interviews, newspapers, etc., that were used to support

your experimental protocols, discussions and conclusions, should be cited in this section.

Important Points to Keep in Mind



Do not confuse the sections of your paper. Pay attention to the difference between the results

and discussion section.

Be clear, concise and complete.

24


Lab 2: Writing a Lab Report




If your results are inconclusive, as are most experiments, say so.

Proof read your report. A lab report is expected to be able to withstand scrutiny.

Do not plagiarize; give credit to all references used.

Experiment 1: Design an experiment

The following experiment is meant to be designed by you! With the beans provided in the kit, you will

design and execute an experiment to test several factors that influence seed germination. Whatever

your experimental design, be sure to include controls and make sure it is reproducible!

Materials

100 beans *You must provide

10 5 x 8in bags

Permanent marker

Ruler

Paper towels*

Water*

Masking tape*




Notes about bean germination:

The time to germination will decrease if you soak the beans overnight

It may take 7‐10 days for the beans to ‘sprout’

Make sure the paper towels remain moist for the duration of your experiment

Procedure

1. Think of 10‐20 variables that may affect seed germination, recording them in Table 3.

2. From your list of variables in Table 3, select three to test. Form a hypothesis for why each affects

seed germination.

3. To germinate the beans, place one folded paper towel, moistened but not soaking wet, into the 5 x

8in bag. Place 10 beans in a horizontal line on the paper towel (between the paper towel and bag).

4. Label each bag with the variable being tested.

25


Lab 2: Writing a Lab Report

Table 3: Variables that may influence seed germination

Variable

Hypothesized Effect

5. Hang each bag vertically using masking tape in the environment you select.

6. Create a table for your data, including title, units, and any other useful information.

7. Select the appropriate type of graph, and report the data you collected.

8. Write a lab report for this experiment in the space provided.

26


Lab 2: Writing a Lab Report

27


Lab 2: Writing a Lab Report

28


Introduction

Lab 3

Data Measurement

29


Lab 3: Data Measurement

Concepts to explore:




The metric system

Converting units

Techniques for obtaining accurate measurements

Introduction

Biology relies heavily on the use of numbers, measurements and calculations. Consequently, scientists

use a universal measuring standard called the metric system. Because the metric system is based on

units of ten, it simplifies making conversions within that system.

The basic units of measurement in the metric system are:

Note: In the table below meters are shown

Gram: when measuring mass.

as an example. The prefixes remain the

Liter: when measuring liquid volume.

same with liter or gram.

Meter: when measuring distance.

Each basic unit can be divided or expanded upon using the following prefixes:

Prefix Abbreviation Multiplier used to convert TO

meters

Nano (n) 10 ‐9 0.000000001 1000000000

Micro (µ) 10 ‐6 0.000001 1000000

Milli (m) 10 ‐3 0.001 1000

Centi (c) 10 ‐2 0.01 100

Deci (d) 10 ‐1 0.1 10

Multiplier used to convert

FROM meters

Prefix Abbreviation Multiplier used to convert TO

meters

Multiplier used to convert

FROM meters

Deka (da) 10 1 10 0.1

Hecto (h) 10 2 100 0.01

Kilo (k) 10 3 1000 0.001

Mega (M) 10 6 1000000 0.000001

Giga (G) 10 9 1000000000 0.000000001

To convert between units, multiply using the conversions above (conversions can also be made by divi‐

31


Lab 3: Data Measurement

sion, though not with this table).

Multiplication Example:


To convert 200 meters (m) to kilometers (km):


multiply 200 m x 0.001 = .2 km

To convert 450 millimeters (mm) to meters (m):

multiply 450 mm x 0 .001 = .45 m

Figure 1: accurate data measurement is

key to reproducible science.

When converting from units less than a meter to greater than a meter (or the other way around), first

convert to a meter and then to the final unit. To convert 40,000 cm to kilometers:




multiply 40,000 cm x 0.01 = 400 m

multiply 400 m x 0.001 = 0.4 km

40,000 cm = 0.4km

Exercise:

1) Convert the following:

3 m = __________ cm

83 m = __________ µm

41,692 m = __________ mm

110 kilometers = __________ m = ____________ mm

3.7 hectometers =_________ m =____________ cm

451,000,000 µm = _________ m = ___________ dam

2) Imagine a field is about 100 meters long. If you run a 5K race how many meters is it? Approximately

how many “fields” does this equate to?

32


Lab 3: Data Measurement

Length, Area, Volume, Mass and Temperature

Length is measured in meters. The area of a square or rectangle is measured by multiplying length (in

meters) by width (in meters). The unit of measurement is m 2 , which reads “meters squared” or “square

meters”. When you see this notation, it is an indication that the measurement is describing area.

Example:

If a box is 12 cm long and 24 cm wide, its area is:

12 cm X 24 cm = 288 cm 2

Volume can be measured by multiplying length (m) by width (m) by height (m). The unit of measurement

is m 3 , which reads “meters cubed” or “cubic meters”. When you see this notation, it is an indication

that the measurement is describing volume.

Example:

If the same box is 4 cm high, its volume would be:

12 cm X 24 cm X 4 cm = 1,152 cm 3

If we wanted to convert this to meters:

1,152 cm 3 X 0.01 = 11.52 m 3

Exercise:

3) Measure the following objects.

A) Your computer screen (in meters)

Length______________

Width ______________

Area _______________

Volume ____________

B) A 100 mL beaker: (in millimeters)

Length _____________

Width _____________

Area ______________

Volume ___________

C) Your lab kit box lid: (in centimeters)

Length _____________

33


Lab 3: Data Measurement

Width _____________

Area _______________

Volume ____________

Mass is the amount of matter an object possesses.

It is the metric systems measurement of weight

and is expressed in grams (g). When using instruments,

such as a scale, there is always a margin of

error. This is a result of either human or mechanical error. Therefore, it is prudent to perform measurements

at least three times to find the average (most precise) measurement.

Exercise:

4) Determine the mass of the objects listed below (in grams). Pay attention to the units. Since you

do not have a metric scale, we will provide you data to work with.

A) Baseball

B) Piece of fruit

Mass (measurement 1): ____.145__kg

Mass (measurement 2): ____145.05_ g

Mass (measurement 3): 145,750.77 mg

Mass (average): __________g

Convert: ___________kg

Mass (measurement 1): ____310____ g

Mass (measurement 2): ___0.318____kg

Mass (measurement 3): __309,143___ mg

Mass (average): __________cg

Convert: ___________ g

Figure 2: Be aware of the margin of error possible

with instruments

Volume is a three dimensional measurement of how space is occupied. Previously we expressed volume

in m 3 (cubic meters). However, the measurement can be reported in units of cubic length or liters.

To convert from one to the other, the conversion 1 cm 3 = 1ml is used.

34


Lab 3: Data Measurement

Example:

To determine the volume of a measurable object, multiply

length x width x height. If a wooden block is 15 cm long, 20

cm wide and 4 cm high, the volume can be found by:

NOTE: When an object is solid

and does not have measurable

sides (i.e. a solid marble), water

displacement can be used to

determine the volume.

Volume = 15 cm x 20 cm x 4 cm

= 1,200 cm 3

= 1,200 ml

= 1.2 L

A graduated cylinder is often used to measure volumes. The graduated cylinder is filled with water and

this initial volume is recorded. The object is added carefully and the new volume is recorded. The difference

of these two volumes is the volume of the object!

Ex: The initial water level in a graduated cylinder is 25.8mL. After an irregularly shaped object is

placed into the cylinder, the water level reads 42.9mL. What is the volume of the irregularly

shaped object?

Answer: 17.1mL or 17.1cm 3

When measuring a liquid there is a certain place

that one must measure ‐ the bottom of the meniscus.

The meniscus is the curved line that a

liquid makes when placed in a narrow container.

When looking for the bottom of the meniscus,

one must look straight at it. When one’s

line of sight is too high, then the reading that is

received is too low. When one’s line of sight is

too low, then the reading received is too high.

35


Lab 3: Data Measurement

Exercise:

Determine the volume of the following objects. If you cannot do so by measuring the dimensions, use

a different technique.

A) The chemical box inside of your kit:

Length: ________ m

Width: _________ m

Height: _________ m

Volume: ________ L

B) Test tube:

Length: _________m

Width: __________ m

Height: __________ m

Volume: _________ L

C) Pick an object from your home. Object:______________.

Length: _________ m

Width: __________ m

Height: __________ m

Volume: _________ L

Exercise

1. If you want to determine the volume of a swimming pool, name two ways you could do this.

2. Measure the volume of a soup bowl from your cupboard. Volume: _________ mL

36


Lab 3: Data Measurement

Temperature is a measure of the amount of heat present in an object. We use the Fahrenheit scale in

the U.S., but the scientific standard is Celsius. In Celsius, water boils at 100 o C and freezes at 0 o C. To

convert between Fahrenheit and Celsius, use the following equation:

o

C = 5/9 ( o F – 32 o )

Example: the human body has a temperature of 98.6 o F:

o C = 5/9 (98.6 o F – 32 o )

o C = 37

Exercise:

1. Convert the following:

121 o F = _______________ o C

32 o F = ________________ o C

0 o F = ________________ o C

77 o F = ________________ o C

2. With your thermometer, measure the temperature of the following objects:

A) Glass of cold tap water: _______________ o C

B) Your kitchen: _______________ o C

C) Inside your freezer: _______________ o C

D) Palm of your hand (wrap your hand around the thermometer, but do not squeeze):

_____________ o C

37


Introduction

Lab 4

Introduction to the Microscope

39


Lab 4: Introduction to the Microscope

Concepts to explore:






Types of microscopes

Parts of a microscope

How to use a microscope

Preparing a wet mount slide

Depth of field

Introduction

Some objects are far too small to be seen with the human eye. However, by using a microscope many

can be viewed in great detail. There are many types of microscopes that range from low–level magnification

(i.e., hand‐held magnification lens) to very high‐power magnification (i.e., an electron microscope).

In the middle of that range lies the light microscope, or for our purposes, the compound light

microscope, which uses multiple lenses.

The compound light microscope (Figure 1) has two sets of lenses:



the ocular lenses (close to your eyes)

the objective lenses (close to the “object” on the stage).

Along with a light source, these lenses work together to magnify the object being viewed. In the case

of the compound light microscope, the total magnification is equal to the magnification power of the

ocular lens multiplied by the magnification power of the objective lens. For example, if the ocular lens

magnifies 10X (this means 10 times) and the objective lens magnifies 10X, the total magnification is

100X.

41


Lab 4: Introduction to the Microscope

Figure 1: A compound microscope can magnify objects that are not visible to the naked eye so that

they can be studied.

Parts of a Compound Light Microscope

Base: The flat support of the microscope.

Light: Illuminates the object being viewed. This can be either in the form of a light source or a

mirror that reflects ambient light onto the image. In the latter case it is important to

be working in an environment with adequate ambient light.

Stage: Supports the slide or other material to be viewed.

Diaphragm: Controls the amount of light allowed on the object.

Stage Clips: Secure the slide in place.

Revolving Nosepiece: Rotates the objective lenses of different magnifications and allows one

of them to be positioned over the slide.

42


Lab 4: Introduction to the Microscope

Arm: Connects the lower base and the upper head of the microscope (also used to carry the

microscope).

Head: Supports both the ocular lens and the revolving nosepiece.

Ocular Lens (eyepiece): The lenses on the microscope typically have a magnification of 10X. If

your microscope has a pointer, which is used to indicate a specific area of the specimen,

it is attached here.

Types of Microscopes

Monocular Microscope: Has a single ocular eyepiece.

Binocular Microscope: Has two ocular eyepieces.

How to Use a Microscope

1. Always carry a microscope with one hand securely around the arm and the other underneath

the base for support.

2. Place the microscope on a table, plug it in, and turn on the light source (or adjust the

mirror as necessary).

Note: When cleaning a microscope, do not use paper towels or cloths as this will

scratch the lens. To preserve the microscope, use only lens paper that will not scratch

the optics.

3. To prevent damage to the lens or slides, always start and end with the scanning power

objective lens (the shortest one) above the light source.

4. Place your slide on the stage and secure it with the stage clips. It is helpful to visually

orient the slide so the object to be viewed is directly in the middle of the opening in

the stage where the light is directed up toward the slide.

5. Turn the course adjustment knob to bring the stage all the way up to the scanning

power objective lens. While looking through the lens, use the course adjustment knob

to slowly lower the stage until the specimen comes into focus.

Note: When using a binocular microscope, adjust the distance between the two

oculars until only one object is seen. Record this distance and set your microscope

to this distance every time you use it. If someone else uses the microscope, the

lenses may be re‐adjusted for their eyes.

6. To adjust the light, open or close the diaphragm located over the light source. When

properly illuminated, the specimen should not be gray or exceptionally bright.

43


Lab 4: Introduction to the Microscope

7. With the object is in general focus, rotate the revolving nosepiece to the low‐power

lens (the next longest). After focusing with the course adjustment knob, switch to the

fine adjustment knob to obtain more precise and greater detail. It may also be necessary

to adjust the light, because more light reduces contrast (sharpness).

8. To become familiar with the mechanical stage knobs around the base of the microscope

(if present), turn one slowly to the right, noting that the image will be moving

toward the left. This image inversion is caused by the lenses.

9. If you need higher magnification, slowly rotate the high‐power lens into place (the next

longest lens). This will bring the tip of the lenses very close to the slide.

10. Make sure the objective lens does not touch the slide.

11. Whenever you use the high‐power lens, only use the fine adjustment knob. If the object

was well focused while viewing with the low‐power lens, very little adjustment

should be necessary.

12. If you cannot bring the object into focus, return to the low‐power lens, focus the object,

and then return to the high‐power lens.

13. When finished, move the revolving nosepiece to the scanning objective lens position

before removing the slide.

How to Prepare a Wet Mount Slide

1. To make a wet mount for a specimen that is not already in liquid, take a clean slide and

place the specimen in the center.

2. Add a drop of water.

Note: For cells that are transparent, it may be necessary to add a small drop of stain as

opposed to water.

3. Carefully add a coverslip by placing one end down and slowly lowering the other end.

Note: If the coverslip is added too quickly, large air bubbles may become trapped which

can cause difficulty viewing the slide. If this happens, gently remove the coverslip, add

another drop of water and try again.

4. Remove excess liquid on the bottom of the slide or around the edges before it is placed

on the microscope to avoid damage to the lens. Just touch a tissue to the edge of the

44


Lab 4: Introduction to the Microscope

coverslip to draw away the water (this is called diffusion and there will be a lab on diffusion

later in the series).

5. If the specimen is already in liquid, place a drop in the middle of the slide and add the

coverslip as you would for a dry specimen.

Experiment 1: Virtual magnification exercise

Materials


“How Big Is It?” demonstration on Student Portal

Note: Review the directions for signing in to the Student Portal at the

beginning of this manual if uncertain how to access this information

Procedure

1. Log into your eScience Student Portal account and locate the “How Big Is It” demonstration

under the Introduction section.

2. Load the animation and beginning with the head of a pin, increase the magnification by

clicking the arrows below the picture. Note the relative sizes of the objects on the pinhead.

3. Be sure to notice the magnification bar on the lower portion of the demonstration that

shows the magnification required to see the objects.

Questions

1. At what magnification do you first notice the ragweed pollen?

45


Lab 4: Introduction to the Microscope

2. Which is bigger, a rhinovirus or E. Coli?

3. Based on the magnification, how many of the E. Coli can fit into the same space as the

head of a pin?

4. About how many red blood cells could fit across the diameter of a human hair (again,

look at the magnification scale)?

46


Biological Processes

Lab 5

The Chemistry of Life

47


Lab 5: Chemistry of Life

Concepts to explore:

Concepts to explore:


Atoms


Acids and bases



Elements

Compounds


The effects of surface area

and volume


Chemical bonds


Molecules/Macromolecules


Energy and metabolism

Introduction

Remember: Mass is the quantity of

matter an object has; weight is the

force produced by gravity acting on

the mass of an object

It is important to have a general understanding of chemistry

before you can begin to understand how living organisms manage

to reproduce, grow, move, eat, and perform a great many

more functions. To begin understanding the myriad of reactions

that occur within a cell, it is important to review the basics of chemistry. Recall that anything that

occupies space and has mass is called matter; all matter is made of atoms.

Atoms are made of a nucleus and two kinds of subatomic particles: electrons (negatively charged particles),

and protons (positively charged particles). Elements are pure substances that are made of only

one type of atom. More than 90% of matter is composed of combinations of just four elements: oxygen,

carbon, hydrogen, and nitrogen. There are over 100 elements known, each with different proper‐

Figure 1: The periodic table of elements categorizes all of the known elements

49


Lab 5: Chemistry of Life

ties. The periodic table has been used to categorize these elements.

In nature, most elements are not found alone; atoms of most elements

combine with the same or different elements to make compounds.

A compound is a mixture of two or more elements in definite

proportions. These atoms are held together by chemical bonds,

bringing them to a stable state. Chemical bonds also store energy.

The two most common bonds are covalent bonds and ionic bonds.

Covalent bonds form when two atoms share electrons. The simplest

part of a substance that retains the properties of that substance

is called a molecule. Ionic bonds form when an atom or

molecule carries an electrical charge, which attracts an atom or

molecule of the opposite charge.

Very large molecules are termed macromolecules. All living organisms

use the same four types of macromolecules for cellular metabolisms

and reproduction. These common biological macromolecules

are proteins, nucleic acids, carbohydrates, and lipids. The

properties they convey are of great importance to cell function, and

you will learn about each in future labs.

Figure 2: Have you ever drank orange

juice right after brushing your

teeth? Yuck! The displeasing taste

is a result of the acid/base reaction

that occurs when a weak acid

(orange juice) mixes with a weak

base (toothpaste).

Chemical reactions take one or more substance and change it to

create a new substance. This requires energy. When chemical

bonds are broken, energy is made available for the reaction to proceed.

Most reactions also require energy to initiate the reaction. This is called the activation energy,

and it differs for each reaction. Catalysts are chemicals that lower the activation energy. You will learn

about biological catalysts called enzymes later in this manual.

Living things require a constant supply of energy. Throughout this manual, you will learn about the reactions

that take place inside of organisms. The sum of these reactions is called metabolism, and is a

general term used to describe the energy require to keep those reactions occurring.

Two important classes of compounds are acids and bases. Both have physical and chemical differences

that can be observed and tested. Acids ionize in water to produce a hydronium ion (H 3 O + ) and bases

dissociate in water to produce a hydroxide ion (OH ‐ ). A compound’s acidity or alkalinity (how basic it is)

can be measured on a scale called pH. The pH of a substance is a measure of the concentration of hydronium

ions. A solution that contains a lot of hydronium ions but few hydroxide ions is considered to

be very acidic. In contrast, a solution that contains many hydroxide ions but few hydronium ions is considered

to be very basic. pH values range from 1‐14, with 1 being highly acidic, 14 highly basic, and 7

neutral.

Have you ever wondered why cells are the size they are? There are many reasons, but one important

one is the surface area to volume ratio. In subsequent labs, you will learn how cells divide once they

50


Lab 5: Chemistry of Life

reach a critical size. Nutrients and oxygen need to diffuse through the cell, and waste needs to diffuse

out of the cell. This must happen quickly for the cell to survive – which happens when the surface area

to volume ratio of the cell is high.

Experiment 1: What household substances are acidic or basic?

There are chemicals, called pH indicators, which change color when they come into contact with an

acid or a base. In the following experiment, you will be using pH paper to determine the pH of various

household substances. The key below indicates the color the paper turns as a function of the pH. In this

way, pH paper allows scientists to determine to what degree a substance is acidic or basic and can provide

an approximate pH value.

Materials

10 1in pH paper strips

5mL Vinegar (


Lab 5: Chemistry of Life

Figure 3: pH paper key

Procedure

1. Find four household substances to test (ex: grape juice, lemon juice, dishwashing liquid, milk,

tomato juice, shampoo, corn starch solution, etc.). You will use the vinegar (acidic) and sodium

bicarbonate (basic) solution provided in your kit as standards.

2. Guess the pH of each substance before testing with pH paper. Record your guesses in Table 1

below.

3. Pour 5mL vinegar into one test tube and 5mL sodium bicarbonate solution in the other.

4. Test each item by dipping one end of the pH paper and record your results in Table 1. (Note:

remember to wear your gloves and goggles when working with chemicals!)

Questions

1. Compare and contrast acids and bases in terms of their H + ion and OH ‐ ion concentrations.

2. Name two acids and two bases you often use.

52


Lab 5: Chemistry of Life

Experiment 2: The effect of surface area and volume

Have you ever wondered why cells don’t grow past a certain size? There is a size limit for cells that

they cannot surpass. Once they reach their maximum size, they divide and form two smaller cells. Why

do they do that? We will look at the importance of surface area to volume ratios in this experiment to

help you understand.

Materials

1 Nutrient Agar Bottle (125 ml)

10mL Bromothymol Blue

Plastic wrap

Pipettes

Vinegar

(1) 250 ml beaker

Rectangular mold

Ruler

Kitchen knife*

Microwave*

Hot pads*

*You must provide

Note: This experiment requires preparation 24 hours in advance.




To determine surface area, multiply Length x Width of each side of the cube and then add them

together. This should be 6 different values added up. For example, if a cube were 3cm x 3cm x

3cm each side would have a L x W = 9 cm 2 . There are six sides in a cube so you would add up the

L x W, in this case 9 cm 2, for all six sides. That would give you a surface area of 54cm 2. . If all the

sides are not the same length, you would determine the length x width for all six sides and add

them up as well.

To determine volume, simply multiple Length x Width x Height.

9 cm 2

To determine surface area to volume ratio, simply divide the surface

area by the volume. 9 cm 2

Procedure

Preparation

1. Begin by removing the cap off the agar solution and placing it in the microwave for 30 seconds.

While in the microwave, watch the solution for boil‐over. If it begins to boil‐over, im‐

53


Lab 5: Chemistry of Life

mediately stop the microwave, but leave the solution inside for two minutes to cool down.

2. After 30 seconds in the microwave, remove the bottle with a hot pad. Screw the lid back onto

the bottle and swirl the solution. If the solution is not completely liquefied, remove the lid

and place the agar bottle into the microwave for 10 second intervals, swirling in between, until

it is completely liquefied. After it is liquefied, let the solution sit for a minute to cool down.

3. Once the agar solution has been removed from the microwave add 10ml of the bromothymol

blue solution to the liquefied agar. Pipette the solution up and down to mix. This should tint

the mixture and allow you to observe a pH change that will occur in subsequent steps.

4. Once the solution is mixed, pour 50ml into the rectangular mold. Cover the container with

plastic wrap and let sit for 24 hours to solidify.

Experiment

1. First, put on safety gloves, safety glasses and apron for safety. Then, check to be sure the agar

has solidified. If it has not, let it sit for another 12 hours.

2. Begin by inverting the rectangular mold, letting the agar block fall onto the underpad.

3. From this block, safely cut out a 1cm x 1cm x 6cm cube. Note: Be sure to measure out the 6cm

first as to avoid any errors.

4. From the remaining agar, safely cut out a 1cm x 1cm x 1cm block. Set the block aside.

5. From the remaining agar, safely cut out a 2cm x 2cm x 2cm block. Set this block with the others.

6. Once all three blocks have been cut, dispose of the scraps of remaining agar. Do not dispose of

the blocks you just cut.

7. Then, measure the surface area, volume and surface area to volume ratio for each cube. Write

those in Table 2.

8. Fill the cleaned 250mL beaker with 150mL of vinegar. Gently place all three blocks into the

vinegar solution.

9. Observe as the blocks begin to change colors. Let them sit in the vinegar for 5 minutes.

10. After 5 minutes, remove the blocks from the vinegar solution. Pour the remaining vinegar solution

down the drain.

11. Gently blot the cubes dry and then safely cut the cubes in half. For each cube, measure the

distance the vinegar diffused into the gelatin cube, as detected by the color change. Do this by

measuring from the outer edge of the cube to the blue rim inside the cube. Record that value

in Table 2.

54


Lab 5: Chemistry of Life

Table 2: Results from surface area to volume experiment

Cube Dimensions Surface Area (cm 2 ) Volume (cm 3 ) Surface Area:Volume Distance of Diffusion

1cm x 1cm x 1cm

2cm x 2cm x 2cm

1cm x 1cm x 6cm

Questions

1. How did the surface area affect the diffusion of the cube? What about the volume? What

about the surface area to volume ratio? Which of these had the greatest affect on the diffusion

of the cube?

2. How does this experiment demonstrate the need for larger cells to divide?

3. For the three cubes shown below, determine their surface area, volume and surface area to

volume ratio. Then, circle the one you believe would be the most efficient and write a summary

stating why.

1.5 cm x 1.5 cm x 1.5 cm

.5 cm x .5 cm x 6 cm

3 cm x 2 cm x 2 cm

55


Biological Processes

Lab 6

Diffusion

57


Lab 6: Diffusion

Concepts to explore:







Diffusion

Rate of diffusion

Direction of diffusion

Concentration gradient

Membrane permeability

Dialysis

Introduction

Molecules are constantly in motion due to the kinetic

energy present in every atom. This energy results in

the net movement of molecules from areas of high

concentration to areas of low concentration, or diffusion

(Figure 1). If uninhibited, this movement will

continue until equilibrium is reached and the molecules

are uniformly distributed.

The rate of diffusion depends on the medium used,

size of the molecule, and polarity of molecule. Because

the medium will not change in a biological system,

the diffusion rate is usually dictated by molecular

characteristics. Small, non‐polar molecules exhibit

a higher rate of diffusion than large, charged

ones.

Figure 1

Diffusion through a semi‐permeable membrane

(lipid bilayer)

The direction of diffusion depends on concentration

gradients, heat and pressure. The concentration gradient is the change of molecular density over a

given area. Temperature and pressure typically remain constant in biological systems, making the concentration

gradient the best indicator of directionality. In general, molecules will move towards areas

of lower concentrations.

A major determinant of diffusion within a biological system is membrane permeability. Cells, as well as

organelles within the cell, are surrounded by selective and differentially permeable membranes. These

membranes control the interaction of the cell and its surrounding environment. Acting as a living gatekeeper,

the membrane allows, slows, or denies access into the cell.

Cellular membranes are composed of two layers of hydrophobic lipids. This lipid bilayer selects for

molecules that can dissolve into the lipid environment and against those that cannot. The ability of a

59


Lab 6: Diffusion

molecule to cross the membrane is also determined

by its size. Small, uncharged molecules pass easily,

while most large or charged molecules are prevented

from passing. Molecules which cannot diffuse across

the membrane may be able to cross through other

regulated gateways.

Hemodialysis is a method of removing toxic

substances from the blood when the kidneys

are unable to do so. It is frequently used for

patients with kidney failure, but may also be

used to quickly remove drugs or poisons in

dangerous situations.

Dialysis is the separation of molecules through diffusion.

A differentially permeable membrane is used to

separate the components of a mixed solution containing

more than one type of molecule. This membrane allows the free passage of water, but limits

the movement of molecules by their size. In one of the following exercises you will dialyze a solution of

glucose and starch to observe membrane permeability.

We will test the rate of diffusion for two dyes. Their different molecular weights (M.W.) will enable you

to observe the effect of molecular size on the rate of diffusion.

Experiment 1: Diffusion through a liquid

Materials

Blue dye (M.W.= 793 g/mol)

Red dye (M.W.= 496 g/mol)

4 Micropipettes

Corn syrup

1 Petri dish (top and bottom)

Ruler

Concepts to explore:

Watch*

Viscous liquid from your cupboard*

* You must provide

IMPORTANT NOTES:

Perform the experiment on only one side of the plate at a time.

Once the syrup has reached the edges of the dish, place the drop of dye on the plate right away.

If you wait too long, the syrup will develop a “skin” and the dyes will not be able to penetrate into

the solution.

Procedure

1. Separate the bottom and top of the petri dish. Place the two dishes over a ruler, making sure

that you can read the markings on the ruler through the dish.

2. Fill the two halves of the petri dish with corn syrup until the entire bottom surface is covered.

60


Lab 6: Diffusion

3. Using a micropipette, place a single drop of blue dye in the middle of the petri dish lid. Note

where the drop fell on the ruler.

4. Measure the diameter of the dye (the distance it has traveled) every 10 seconds for a total of 2

minutes. Record your measurements in the following tables (Tables 1, 2, and 3) and graph

your results (distance traveled on the y axis, time on the x axis).

5. Repeat this process using the red dye in the bottom half of the petri dish.

6. After you have recorded your results, clean out the petri dish halves.

7. Choose one other liquid from your cupboard and repeat steps 2‐5 using your chosen material

in place of the corn syrup.

8. Record your results in Table 1.

Table 1:Diffusion Through Liquids

Corn Syrup

Material:______________

Time (sec) Blue Dye Red Dye Blue Dye Red Dye

10

20

30

40

50

60

70

80

90

100

110

120

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Lab 6: Diffusion

Table 2: Diffusion Rate of Dyes in Corn Syrup

Dye Name Molecular Weight Total Distance Traveled (mm) Speed of Diffusion (mm/hr)*

Blue Dye

Red Dye

Table 3: Diffusion Rate of Dyes in ________________

Dye Name Molecular Weight Total Distance Traveled (mm) Speed of Diffusion (mm/hr)*

Blue Dye

Red Dye

*multiply total distance traveled by 30 to get the hourly diffusion rate

Graphs (don’t forget to label the axes!):

62


Lab 6: Diffusion

Questions

1. Which dye diffused the fastest in corn syrup? In your chosen material?

2. Does the rate of diffusion correspond with the molecular weight of the dye?

3. Does the rate of diffusion change over time? How might this affect your calculated diffusion

rate compared to the actual diffusion rate?

4. Cells receive vital nutrients and rid toxic waste with the help of the circulatory system. What is

the critical distance a cell must maintain from a capillary (the point of nutrient/waste exchange)

in order to survive? Explain the role diffusion plays in this process.

5. Describe why the medium the dyes diffuse through can affect the rate of diffusion. How does

this relate to nutrient transport into cells?

63


Lab 6: Diffusion

Experiment 2: Concentration gradients and membrane permeability

We will dialyze a solution of glucose and starch to observe:

The directional movement of glucose and starch.

The effect of a selectively permeable membrane.

In this lab, we will be using an indicator to test for the presence of starch and glucose. An indicator is a

substance that changes color when in the presence of the substance it indicates

Materials

Glucose solution

(M.W.=180g/mol)

1% Starch solution

(M.W.=>15,000g/mol)

Iodine‐Potassium iodide (IKI)/Lugol’s

Solution

4 Glucose test strips

15cm Dialysis tubing**

4 100mL Beakers

4 Small rubber bands

7 Graduated pipettes

Water*

Watch*

*You must provide

** Cut to exact length




Note:

You will need dialysis tubing in subsequent experiments, so be sure to cut the amount

specified in the directions.

Dialysis tubing can be rinsed and used again if you make a mistake.

Dialysis tubing must be soaked in water before you will be able to open it up to create

the dialysis “bag”. Follow the directions for the experiment, beginning with soaking the

tubing in a beaker of water. Then, place the dialysis tubing between your thumb and

forefinger and rub the two digits together in a shearing manner. This should open up

the "tube" so you can fill it with the different solutions.

Procedure

1. Fill one 100mL beaker with 50mL water and submerge the dialysis tube for 10 minutes. Fill a

second beaker with 80mL water (this is the one you will put the filled dialysis bag into in Step

9).

2. After the ten minutes have passed, remove the dialysis tube and close one end by folding over

3cm of one end (bottom). Fold it again and secure with a rubber band (use two if necessary).

3. Make sure the closed end will not allow a solution to leak out. You can test this by adding a few

drops of water and looking for leakage. Pour the water out before continuing.

64


Lab 6: Diffusion

Attention!

Do not allow the open end of the bag to fall

into the beaker. If it does, remove the

tube and rinse thoroughly with water

before refilling with a starch/glucose

solution and replacing it in the beaker.

Figure 2: Experimental set‐up

4. Use a graduated pipette to add 5ml of glucose solution to a

third beaker and label it “Dialysis bag solution”. Using another

graduated pipette, add 5mL of starch solution to the

same beaker. Mix by pipetting the solutions up and down

the pipette six times.

Indicator Reagents

IKI Solution:

Yellow = no starch

5. Transfer 8mL of the dialysis bag solution (glucose and starch) Purple/Black = starch

into the prepared dialysis bag. The remaining 2mL will serve

Glucose Test Strip:

as a sample to test for the presence of glucose and starch (to

act as a control and show that both glucose and starch were Yellow = no glucose

present in the solution poured into the dialysis bag).

Green = glucose

6. Label the last (fourth) beaker “Beaker solution”, and using a

clean pipette, transfer a sample of 2mL of the water in the

second beaker to this beaker (to act as another control to show that the water the dialysis bag

is placed in does not contain starch or glucose).

7. Test for the presence of glucose by dipping one glucose test strip into the dialysis bag solution

sample (third beaker) and another strip into the beaker solution sample (fourth beaker). Wait 1

minute, then observe the color of the test strip. Record your results in the following tables

(Tables 4 and 5).

8. Next, add a few drops of IKI solution into both sample beakers (the third and fourth beakers).

Record your observations in Tables 4 and 5.

9. Place the filled dialysis tube into the second beaker filled with 80mL of water with the open

end draped over the edge of the beaker as shown below.

10. Use a pipette to add 2ml of IKI solution to the beaker water. Record the initial color of both

the beaker water and the solution in the dialysis tube in the table below (Table 4).

11. After the solution has diffused for 60 minutes, remove the dialysis tube from the beaker.

65


Lab 6: Diffusion

Beaker

12. Again, test for the presence of glucose by dipping one glucose test strip into the dialysis bag

directly and another strip into the beaker solution. Again, wait one minute before reading the

results of the test strip. Record your results for the presence of glucose and starch in the following

Tables 4 and 5.

Dialysis tube

Table 4: Starch Diffusion

Initial color Starch present? Final color Starch present?

Table 5: Glucose Diffusion

Beaker

Dialysis tube

Present/Absent

Before Dialysis

Present/Absent

After Dialysis

Questions

1. Which substance crossed the dialysis membrane? What evidence from your results proves

this?

2. What molecules remained inside of the dialysis bag?

3. Of the substances that diffused through the bag, did all of the molecules diffuse out?

4. Does the dialysis bag or the beaker contain more starch? What about glucose?

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Lab 6: Diffusion

5. Is the bag hypotonic with regards to the IKI solution, or the beaker? What about the starch

solution?

6. What results would you expect if the experiment started with glucose and IKI Solution inside

of the bag, and starch and water in the beaker? Why?

7. Draw a diagram of this set up. Use arrows to depict the movement of each substance in the

dialysis bag and the beaker.

8. What type of membrane does the dialysis tubing represent? Give an example of this type of

membrane that can be found inside the body.

9. How does the glucose concentration affect diffusion rate?

67


Biological Processes

Lab 7

Osmosis

69


Lab 7: Osmosis

Concepts to explore:






Osmosis

Hypertonic

Hypotonic

Isotonic

Osmotic pressure

Introduction

A major determinant of diffusion in a biological system is membrane permeability. Small, uncharged

molecules pass through cellular membranes easily, while most and/or charged molecules cannot pass

through the membrane.

The movement of water across a selectively permeable membrane, like the plasma membrane of the

cell, is called osmosis. Osmosis occurs when a membrane separates solutions of different concentrations.

The membrane allows the solvent to pass through, but not the solutes. Ultimately, membrane

selectivity and the movement of water in and out of the cell regulates the concentration of intracellular

material. Remember, a solution contains two or more substances (solutes) that have been dissolved

by a solvent. In the context of a cell, the intracellular and extracellular fluids are the solvents which

contain dissolved material (solutes). As solute concentration increases, solvent concentration decreases.

Tonicity is a relative term used to describe osmotic pressure of two solutions separated by a semipermeable

membrane. It is influenced by solutes that cannot cross the semipermeable membrane (solutes

that do cross the membrane will always move to achieve equal concentrations on both sides of the

membrane). Thus, tonicity determines the net direction of movement of water molecules.

Figure 1 The three types of tonicity

71


Lab 7: Osmosis

There are three types of tonicity (Figure 1) —remember that it is a relative term used to compare one

solution to another:




A hypertonic solution contains a greater concentration of solutes unable to cross the membrane

compared to the solution on the other side of the membrane.

A hypotonic solution contains a lower concentration of solutes unable to cross the membrane

compared to the solution on the other side of the membrane.

An isotonic solution has an equal concentration of impermeable solutes on both sides of the

semi‐permeable membrane.

When osmosis takes place, water flows from hypotonic solutions to hypertonic solutions, until the solutions

become isotonic. In most biological systems, cells are hypertonic and extracellular water flows

into them. If placed in pure water, they will burst (lyse) as a result of the increased pressure on the

membrane from the additional water that diffused into the cell.

Osmotic pressure (the force required to prevent osmosis) is directly correlated with tonicity (higher

tonicity causes an increase in osmotic pressure). Some cells, such as plant cells, have specialized structures

that regulate osmotic pressure and prevent lysis.

Experiment 1: Direction and concentration gradients

In this experiment, we will investigate the effect of solute concentration on osmosis. A semi‐permeable

membrane (dialysis tubing) and sucrose will create an osmotic environment similar to that of a cell.

Using different concentrations of sucrose (which is unable to cross the membrane) will allow us to examine

the net movement of water across the membrane.

Materials

30% Sucrose solution

4 15cm Pieces dialysis tubing**

3 250mL Beakers

8 Rubber bands

10mL Graduated cylinder

Blue, red, yellow, green beads

Concepts to explore:

Water*

Watch*

*You must provide

**Cut to exact length

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Lab 7: Osmosis




Note:

You may need dialysis tubing in subsequent experiments, so be sure to cut the amount

specified in the directions.

Dialysis tubing can be rinsed and used again if you make a mistake.

Dialysis tubing must be soaked in water before you will be able to open it up to create

the dialysis “bag”. Follow the directions for the experiment, beginning with soaking the

tubing in a beaker of water. Then, place the dialysis tubing between your thumb and

forefinger and rub the two digits together in a shearing manner. This should open up

the "tube" so you can fill it with the different solutions.

Procedure

1. Submerge the four pieces of dialysis tubing into a 250 mL beaker

filled with 100 ml of water for at least 10 minutes.

2. After 10 minutes, remove one piece of tubing from the beaker.

On one end (not the whole tube), gently twirl the tubing into a

long, thin cylindrical piece that is able to fit into the hole of the

yellow bead.

3. Insert the long cylindrical end of the tube into the center hole in

the yellow bead. Once it is through, pull the cylindrical end until

there is about 1.5 to 2cm of tubing extending beyond the bead

Figure 2: Fold the bag until you

have a piece narrow enough to

be threaded through the bead.

4. Take the extra tubing you just pulled through the bead and fold it back over the bead, towards

the remaining, non folded tube. Place a rubber band above the bead and around the extra

tubing as to be sure no solution can leak out of the tube (see Figure 2).

Figure 3: Beads help to secure the ends of the dialysis bags and identify each one.

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Lab 7: Osmosis


To test that no solution can leak out, add a few drops of water and look for water leakage.

Make sure you pour the water out before continuing to the next step.

5. Repeat steps 2‐4 with the three remaining dialysis tubes, using each of the three remaining

bead colors (Figure 3).

6. Table 1 provides a distinction as to what bead belongs to which tube. Using a 10mL graduated

cylinder, measure and fill the appropriate dialysis bag with the designated concentration of sucrose

solution (3%, 15% or 30%) by adding the volumes of sucrose and water listed in the table

below.

Table 1: How to make a serial dilution of sucrose

Bead Color Bag Number Stock Sucrose Solution Water

Yellow Bag #1: 30% sucrose 10mLs 0mLs

Red Bag #2: 15% sucrose 5mLs 5mLs

Blue Bag #3: 3% sucrose 1mL 9mLs

Green Bag #4: 3% sucrose 1mL 9mLs

7. Rinse the outside of the bags with water to remove any remaining sucrose.

8. Pour 150mL of the stock sucrose solution (30%) into the 250mL beaker (beaker #1). Using the

graduated cylinder, measure 20mLs of the stock sucrose solution and 180mL of water to create

a 3% sucrose solution and place it into the 250mL beaker (beaker #2).

9. Place bags #1‐3 (red, blue, yellow) into beaker 2 and bag #4 (green) into beaker 1 (Figure 4).

Figure 4: The dialysis

bags are filled with

varying concentrations

of sucrose solution

and placed in one

of two beakers.

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Lab 7: Osmosis

10. In Table 2, predict whether water will flow in or out of each dialysis bag.

11. Allow the bags to sit for one hour. While waiting, dump out the water in the 250 mL beaker

that was used to soak the dialysis tubing in step 1. We will use this in the last part of the experiment.

12. After allowing the bags to sit for one hour, remove them from the beakers.

13. Carefully open the bags, noting that often times the tops may need to be cut as they tend to

dry out. Measure the solution volumes of each dialysis bag using the empty 250 ml beaker.

Record your data in Table 2.

Table 2: Water movement

Initial Volume Sucrose % Prediction: Will water move in or out? Final Volume

Bag#1 10mL

Bag #2 10mL

Bag #3 10mL

Bag #4 10mL

Questions

1. For each of the bags, identify whether the solution inside was hypertonic, hypotonic or

isotonic in comparison to the beaker solution it was placed in.

2. Which bag increased the most in volume? Why?

3. What does this tell you about the relative tonicity between the contents of the bag and the

solution in the beaker?

4. What would happen if bag 1 is placed in a beaker of distilled water?

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Lab 7: Osmosis

5. Osmosis is how excess salts that accumulate in cells are transferred to the blood stream so

they can be removed from the body. Explain how you think this process works in terms of

tonicity.

Experiment 2: Tonicity and the plant cell

Plant cells are able to generate osmotic pressure while other cells cannot. This is due to specialized

plant structures such as the cell wall which prevent lysis caused by osmosis. By taking advantage of this

system, you will be able to look at the effects of tonicity in a biological system.

Materials

20% Sodium chloride (NaCl) solution

Several types of potatoes (e.g. russet,

Yukon, yams)*

2 Plastic test tubes

100mL Graduated cylinder

Water*

2 Pipettes

Knife for cutting*

Ruler

Paper towel*

Watch*

Permanent marker

*You must provide

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Lab 7: Osmosis

Procedure

1. Label two test tubes (A and B) for each type of potato you will be testing. Be sure to write the

type of potato on the tube as well. Fill in the types of potato used in this experiment in the

“Type of Potato” column of Table 3.

2. Carefully cut two strips of each type of potato on a cutting board. The strips should be as close

to 10cm long and 1.5cm wide as you can cut them.

Note: In the next step, you will account for any variability by measuring the volume of water

displaced when submerged into a beaker containing a known volume of water.

3. Fill the 100mL graduated cylinder with 50mL of water. Place one strip of the first type of potato

(Sample A) into the graduated cylinder and record the amount of water it displaced in the

“Initial Displacement” column of Table 3 in the row corresponding with the sample tested.

Note: Displacement is a measurement of change and is calculated by subtracting the original

volume (50ml) from the final volume that you read after the potato is added to the 50mls of

water. (eg. 57mL—50mL = 7mL)

4. Remove Sample A from the graduated cylinder and, if any water was lost, fill the graduated

cylinder up again with 50mL of water. Place Sample A into the corresponding test tube.

5. Place the second strip of the same type of potato (sample B) into the graduated cylinder and,

again, record the amount of water it displaced in the “Initial Displacement” column of Table 3

row corresponding with the sample tested.

6. Remove Sample B from the graduated cylinder and, if any water was lost, fill the graduated

cylinder up again with 50mL of water. Place Sample B into the corresponding test tube.

7. Repeat Steps 3‐6 for each type of potato you will be testing.

8. Using the plastic dropper, add water to each of the test tubes with the A samples in them until

the water covers the potato strip. In a similar manner, add the 20% Sodium Chloride (NaCl)

solution to each of the test tubes containing the B samples.

Note: Make sure your test tubes are upright during the experiment. It may be useful to use the

test tube rack provided.

9. After an hour, drain the liquid from the test tubes containing your samples.

10. Fill the 100mL graduated cylinder with 50mL of water. Repeat Steps 3‐6 for each sample and

record the displacement in the “Final Displacement” column of Table 3 row corresponding with

the sample tested.

11. Complete the last column of Table 3 by subtracting the initial displacement from the final displacement.

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Lab 7: Osmosis

Table 3: Amount of water displaced by potato samples before and after experiment

Type of Potato

Sample

Initial Displacement

(mL)

Final Displacement

(mL)

Net Change=

Final Displacement

‐ Initial Displacement

A

B

A

B

A

B

Questions

1. What is measured when looking at the net change in displacement of the potato samples?

2. Different types of potatoes have varying natural sugar concentrations. Explain how this

may influence the experiment.

3. Based on the data from this experiment, hypothesize which potato has the highest natural

sugar concentration. Explain your reasoning.

4. What was the texture and your observation of the potato samples prior to the experiment?

After? Did it vary by type of potato?

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Lab 7: Osmosis

5. Would this experiment work with other plant cells? What about animal cells? Why or why

not?

6. From what you know of tonicity, what can you say about the plant cells and the solutions

in the test tubes?

7. What do your results show about the concentration of the cytoplasm in the potato cells at

the start of the experiment?

8. If the potato is allowed to dehydrate by sitting in open air, would the potato cells be more

likely to absorb more or less water? Explain.

9. Could this experiment be performed with other solutions (i.e., sugar)? Why? Design an experiment

to test the effect of different solutions (be sure to include controls) on tonicity in

plant cells. Note: Use something other than a potato in your experiment!

79


Biological Processes

Lab 8

Respiration

81


Lab 8: Respiration

Concepts to explore:





Cellular energy

Respiration

Anaerobic respiration

Aerobic respiration

Introduction

ATP is the energy currency of the cell. It is produced through a process called respiration.

The energy molecules (ATP) generated through respiration, are available to fuel the processes of the

cell as needed. When ATP levels become too low a special protein signals the cell to begin respiration.

As long as all the critical components for the reaction are available, this cycle provides a constant

source of energy for the cell.

Respiration harvests biological energy from fuel molecules, such as

carbohydrates, and stores it as ATP. Together with oxygen, the cell

converts carbohydrates to carbon dioxide, water and most importantly

energy. Respiration is a controlled, multistep process which

slowly releases the energy stored in glucose and converts it to ATP.

If all of this energy from glucose were released at once, most

would be lost as heat and light.

Carbohydrates contain high energy bonds that, when broken, release

electrons. The first stage, glycolysis, breaks carbohydrates

(glucose) into pyruvate molecules. Though the bonds holding pyruvate

together contain a great deal of potential energy, this step

yields little energy.

Yeast has been used to make

leavened bread for centuries.

When yeast undergoes

fermentation, CO 2 is trapped

between gluten and causes the

bread to rise. Ethanol, another

byproduct of yeast fermentation,

generates the alcohol content in

beer, and the CO 2 provides

effervescence. What ingredients

must be present in order for this

process to occur?

Glycolysis occurs with or without oxygen and takes place in the cytoplasm outside the mitochondria.

Interestingly, it is a pathway found in all living things.

C 6 H 12 O 6 + 6O 2 6CO 2 + 6H 2 O + energy

glucose oxygen carbon dioxide water

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Lab 8: Respiration

Membrane

Anaerobic

2 ATP

Aerobic

in mitochondria

6 Carbon Compound

Glucose

4 ADP

Oxidation

3 Carbon Compound

Pyruvate

4 ATP

+ O

2

CO 2

+ H2O

Yeast fermentation

2 ATP

Homolactic

fermentation

Alcohol +

CO 2

Lactic

Acid

34 ADP

34 ATP

Cytosol

Mitochondrion

Figure 1: Aerobic Respiration

Aerobic respiration takes place in the mitochondria (a specialized organelle) of the cell and uses oxygen

as the final electron acceptor in the electron transport pathway. Pyruvate is oxidized to generate energy.

Special molecules shuttle electrons to the ATP production site. Since oxygen has a very high affinity

for electrons, aerobic respiration is the most efficient means of producing ATP (36 per reaction).

Anaerobic respiration takes place in the cytoplasm

of the cell and uses other, less efficient,

molecules to transport electrons.

If the final transfer molecule is organic (contains

a carbon), the process is called fermentation.

Fermentation is an anaerobic process that reduce

regenerate NAD+ so that glycolysis can

continue. Because it cannot fully break down

the glucose molecule, fermentation is far less

efficient than aerobic respiration, generating

only two ATP molecules.

During physical activity, cells require more energy.

As long as enough oxygen can be delivered

to cells, aerobic respiration dominates.

When energy consumption exceeds the oxygen

supply, anaerobic respiration starts. Lactic acid is

a byproduct, and is what causes muscle soreness

after a hard workout!

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Lab 8: Respiration

Experiment 1: Fermentation by yeast

Yeast cells produce ethanol and CO 2 during fermentation. We will measure the production of CO 2 to

determine the rate of anaerobic respiration in the presence of different carbohydrates.

Note: Sucrose (a disaccharide) is made up of glucose and fructose. Glucose is a monosaccharide.

Materials

5 Respirometers (Figure 2)

1% Glucose solution

1% Sucrose solution

Equal, Splenda, and sugar

packets

1 Yeast packet

4 250mL Beakers

100mL Graduated cylinder

Warm water*

Pipettes

Watch or timer*

Permanent marker

Ruler

Measuring spoon

*You must provide

Figure 2: To make a respirometer, obtain two test tubes

that fit into each other – one small plastic test tube and

one large glass test tube for each respirometer.

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Lab 8: Respiration

Procedure

1. Completely fill the smallest tube with water and invert the larger tube over it. Push the small

tube up (into the larger tube) until the top connects with the bottom of the inverted tube. Invert

the respirometer so that the larger tube is upright (there should be a small bubble at the

top of the internal tube). Repeat this several times as practice – strive for the smallest bubble

possible. When you feel comfortable with this technique, empty the test tube and continue

with this experiment.

2. Mix 1/4 tsp of yeast into 175mL of warm (40‐43°C) water in a 250mL beaker. Stir until dissolved.

Note: Make sure the yeast solution is stirred before each test tube is filled.

3. Label both the big and small test tubes 1‐5.

4. In a 250 ml beaker, mix the 1 gram packet of Equal with 100mL of water. In another 250 mL,

mix the 1 gram packet of Splenda with 100mL of water. In another 250mL beaker, mix HALF

of the 4 gram packet of sugar with 200mL of water.

5. Fill the smaller test tubes with 15mL solution as follows:

Tube 1: 1% glucose solution

Tube 2: 1% sucrose solution

Tube 3: 1% Equal solution

Tube 4: 1% Splenda solution

Tube 5: 1% sugar solution

Note: A good practice in the lab is to rinse the graduated cylinder between each use.

6. Then, fill each tube to the top with the yeast solution.

7. Slide the corresponding larger tube over the small tube and invert it as practiced. This will mix

the yeast and sugar solutions.

8. Place respirometers in the test tube rack, and measure the initial air space in the rounded bottom

of the internal tube. Record these values in the Table 1.

9. Allow the test tubes to sit in a warm place (~37˚C) for one hour. A few suggestions are: a sunny

windowsill, atop (not in!) a warm oven heated to 200˚F, under a very bright (warm) light, etc.

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Lab 8: Respiration

10. At the end of the respiration period, measure the air space in the internal tubes, and record

in Table 1.

Table 1: Gas production

Tube Initial gas height (mm) Final gas height (mm) Net Change

1

2

3

4

5

Questions

1. Hypothesize why some substances were not metabolized, while others were. Research the

chemical formula of Equal and Splenda and explain how it would affect respiration.

2. If you have evidence of respiration, identify the gas that was produced. Suggest two methods

for positively identifying this gas.

3. How do the results of this experiment relate to the role yeast plays in baking?

4. What would you expect to see if the yeasts’ metabolism was slowed down? How could this be

done?

5. Indicate sources of error and suggest improvement.

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Lab 8: Respiration

6. Optional: Using the leftover yeast, test some other carbohydrates (e.g. brown sugar, molasses)

from your cupboard to see if the rates of respiration differ. Report your experimental procedure

and results below.

Experiment 2: Aerobic respiration in beans

We will evaluate respiration in beans by comparing carbon dioxide production between germinated

and non‐germinated beans. As shown in the balanced equation for cellular respiration, one of the byproducts

is CO 2 (carbon dioxide):

C 6 H 12 O 6 + 6H 2 O + 6O 2 energy + 6CO 2 +12H 2 O

We will use a carbon dioxide indicator ( bromothymol blue) to show oxygen is being consumed and

carbon dioxide is being released by the beans. Bromothymol blue is an indicator that turns yellow in

acidic conditions, green when it is neutral and blue when it is in basic conditions. When carbon dioxide

dissolves in water, carbonic acid is formed by the reaction:

H 2 O + CO 2 H 2 CO 3

resulting in the formation of this weak acid. If an indicator such as bromothymol blue is present, what

do you think would happen? (Hint—what color would the indicator change to?)

Materials

100 Pinto beans

100 Kidney beans

6 250mL beakers

Paper towels*

6 Measuring cups (small white paper

cups)

Pipette

24mL Bromothymol blue solution

Parafilm

6 Rubber bands

Water*

*You must provide

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Lab 8: Respiration

Procedure

Figure 3: Beaker set‐up

1. Fill two beakers with 200ml water.

2. Soak 50 pinto beans in Beaker 1P and 50 kidney beans in

Beaker 1K for 24 hours.

3. Empty the water from beakers 1P and 1K.

4. Pour the soaked beans onto paper towels, keeping them

separated.

5. Label the remaining beakers: Beakers 2P, 3P, Beaker 2K, and

3K.

6. Place several layers of moist paper towels at the bottom of

the 250ml beakers.

4. Place 50 pre‐soaked pinto beans into Beaker 1P, 50 control pinto beans in Beaker 2P, and zero

beans in Beaker 3P.

5. Place 50 pre‐soaked kidney beans into Beaker 1K, 50 control kidney beans in Beaker 2K, and

zero beans in Beaker 3K.

6. Dispense 4ml of bromothymol blue solution into each of the three measuring cups, and place

the measuring cup inside each beaker (Figure 3).

7. Stretch Parafilm across the top of each beaker. Secure with a rubber band to create an airtight

seal.

Note: If your Parafilm breaks, plastic wrap can also be used.

7. Place the beakers on a shelf or table, and let sit undisturbed at room temperature.

8. Observe the jars at 30 minute intervals for three hours, and record any color change of the bromothymol

blue in Tables 2 and 3.

9. Let the beans and the jar sit overnight. Record your observation in Tables 2 and 3.

Table 2: Bromothymol blue color change over time for pinto bean experiment

Time

Beaker with pre‐soaked

beans

Beaker with un‐soaked beans

Beaker with no beans

0 min

30 min

60 min

90 min

120 min

150 min

180 min

24 hours

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Lab 8: Respiration

Table 3: Bromothymol blue color change over time for kidney bean experiment

Time

Beaker with pre‐soaked

beans

Beaker with un‐soaked beans

Beaker with no beans

0 min

30 min

60 min

90 min

120 min

150 min

180 min

24 hours

Questions

1. What evidence do you have to prove cellular respiration occurred in beans? Explain.

2. Were there differences in the rates of respiration in pinto beans vs. kidney beans? If so, why?

3. If this experiment were conducted at 0°C, what difference would you see in the rate of respiration?

Why?

4. What is the mechanism driving the bromothymol blue solution color change?

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Lab 8: Respiration

5. What are the controls in this experiment, and what variables do they eliminate? Why is it important

to have a control for this experiment?

6. Would you expect to find CO 2 in your breath? Why?

7. What effect would large changes of temperature (e.g., 37°C vs. 45°C) have on respiration in

beans? Design an experiment to test your hypothesis, complete with controls.

91


Biological Processes

Lab 9

Enzymes

93


Lab 9: Enzymes

Concepts to explore:

Concepts to explore:


Enzymes


Activators


Selectivity


Inhibitors


Catalysts


Activation energy


Activation site


Reaction rates

Introduction

Enzymes are specialized proteins that serve as biological catalysts to decrease the activation energy

normally needed for a reaction to occur. This means the reaction rate is up to millions of times faster

than it would be without the enzyme. Most biochemical reactions require enzymes for them to occur

at fast enough rates to be useful. Typical nomenclature for enzymes follows the pattern using the

name of the substrate or the chemical reaction it catalyzes, and ends with “‐ase”, e.g. catalase, amylase.

(In other words, any time you see a word end in “ase” you know it is an enzyme).

Figure 1: The specificity of enzymes is controlled by their lock and key fit with a specific

substrate.

Enzymes are extremely selective, and are often described as having a “lock and key” fit (Figure 1).

Their shape determines which substrates they bind and interact with. The activation site

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Lab 9: Enzymes

is the pocket where the substrate attaches and where the reaction occurs. After the enzyme/substrate

complex forms and catalysis occurs, the “new” substrate is released from the active site, and the enzyme

can repeat the process. Enzymes levels are not reduced or altered during the reaction. This

means they are efficient and can be used repeatedly.

Enzymes determine the rate at which the reaction occurs (not how it occurs). Their activity is affected

by temperature, pH, enzyme and substrate concentration, and other chemicals that may be present

(such as salts, which can change the protein structure).

Variations in temperature and alkalinity can change the shape of the proteins, such as enzymes, which

makes them inactive (they can no longer bind to their substrate). The pH can alter charge of the protein,

once again changing its shape and rendering them inactive.

The concentrations of both the enzyme and substrate determine the reaction rate (Figure 2). Remember

that high reaction rates do not always translate into rapid time of completion (it also depends on

the amount of substrate!).

Figure 2: Substrate Saturation Curve

Activators are chemicals that bind to the active site of the enzyme

and help it to bind to the substrate. They are sometimes

called cofactors or organic coenzymes.

Inhibitors are chemicals that interfere with the binding of the

substrate to the enzyme. There are two types:

Many drugs and poisons are enzyme

inhibitors. For example, aspirin

inhibits an enzyme that leads to

inflammation.

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Lab 9: Enzymes



Competitive (can be replaced by the substrate)

Non‐competitive (not removed by the substrate)

Normal cellular processes produce toxic substances (waste) such as hydrogen peroxide and free radicals

that if not eliminated, will kill the cell. Luckily, yeast and other organisms (including humans) have

an enzyme called catalase that breaks down hydrogen peroxide into oxygen and water, both harmless

to cells.

Experiment 1: Effect of enzyme concentration

Yeast cells contain catalase. The effect of catalase can be seen when yeast is combined with hydrogen

peroxide (Catalase: 2H 2 O 2 ─› 2 H 2 O + O 2 ). In this lab you will examine the effects of enzyme (catalase)

concentration based on the amount of oxygen produced.

Materials

Yeast

Measuring Spoon

3 Test tubes

3 100mL Beakers

Hydrogen peroxide

10 ml Graduated cylinder

Permanent marker

Ruler

String*

3 Balloons

Watch*

*You must provide

Procedure

1. Label three test tubes 1, 2, and 3

with a permanent marker.

2. Fill each tube with 10mL hydrogen

peroxide.

3. Label three beaker A, B, C.

4. Add 1/2 teaspoon yeast (1 g) to 100

ml warm water (30‐35°C) in Beaker

A. Mix well by pipetting.

Figure 3: When catalsae is added to hydrogen peroxide, oxygen

is released.

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Lab 9: Enzymes

5. Make a serial dilution of yeast solution by adding 10mL yeast solution from Beaker A and transfer

to Beaker B. Add 90 mL warm water (30‐35°C) to Beaker B. Mix well by pipetting.

6. Take 10mL yeast solution from Beaker B and transfer to Beaker C. Add 90 mL warm water (30‐

35°C) to Beaker C. Mix well by pipetting.

7. Into the first test tube, add 5mL from Beaker A.

8. Quickly attach a balloon to the top of the test tube so that it will fill with the oxygen produced

by the enzyme reaction. It is important to execute this step quickly so that every bit of gas produced

will be captured.

9. Repeat this procedure by transferring 5ml from Beaker B into test tube 2, attaching the balloon,

and transferring 5 mL from Beaker C into test tube 3 and immediately placing the balloon

on top.

10. Swirl each tube to mix, and wait 30 seconds.

11. Wrap the string around the center of the balloon to measure the circumference. Measure the

length of string with a ruler. Record measurements in Table 1 below.

12. Repeat step 11 for the remaining balloons.

Table 1: Effect of enzyme concentration on the production of gas

Tube Amount of yeast Balloon circumference (cm)

1 0.05g

2 0.005g

3 0.0005g

Questions

1. What is the enzyme in this experiment? What is the substrate?

2. Did you notice a difference in the rate of reaction in the tubes with different concentrations of

enzymes? Why or why not?

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Lab 9: Enzymes

3. What was the effect of using less enzyme on your experiment?

4. Do you expect more enzyme activity if the substrate concentration is increased or decreased?

Draw a graph to illustrate this relationship.

5. Hydrogen peroxide is toxic to cells, yet is a common byproduct of the reactions that occur inside

the body. How can this compound by changed to become non‐toxic (Hint: Look at the

chemical formula of hydrogen peroxide)?

Experiment 2: Effect of temperature on enzyme activity

This experiment looks at the effect of temperature on enzyme activity.

Materials

Yeast

Measuring spoon

3 Test tubes

Hydrogen peroxide

10 mL Graduated cylinder

3 Balloons

2 Beakers

Hot water bath*

Permanent marker

Ruler

String*

Watch*

Thermometer

*You must provide

Procedure

1. With a permanent marker, label test tubes 1, 2, and 3.

2. Fill each tube with 10mL hydrogen peroxide.

3. Place tube 1 in the refrigerator, leave tube 2 at room temperature, and place tube 3 in the hot

water bath (>85°C).

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Lab 9: Enzymes

4. Record the temperatures of each condition in the table below. Let tubes sit for 15 minutes.

5. After the elapsed time, remove tubes from the refrigerator and the boiling water bath.

6. Add 1/4 teaspoon of yeast to the refrigerated test tube.

7. Quickly attach a balloon to the top of the test tube so that it will fill with the oxygen produced

by the enzyme reaction. It is important to execute this step quickly so that every bit of gas produced

will be captured.

8. Repeat steps 6‐7 for the test tube in the hot water bath, then the room temperature test tube.

9. Swirl each tube to mix, and wait 30 seconds.

10. Wrap the string around the center of the balloon to measure the circumference. Measure the

length of string with a ruler. Record measurements in the Table 2 below.

11. Repeat step 10 for the remaining balloons.

Table 2: Effect of temperature on the production of gas

Tube Temperature ˚C Balloon circumference (cm)

Refrigerator

Room temperature

Hot water

Questions

1. What is the enzyme in this experiment? What is the substrate?

2. How does temperature affect enzyme function?

3. Do plants and animals have an enzyme that breaks down hydrogen peroxide? How could you

test this?

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Lab 9: Enzymes

4. How can enzyme activity be increased?

5. Design an experiment to determine the optimal temperature for enzyme function, complete

with controls. Where would you find the enzymes for this experiment? What substrate would

you use?

6. Draw a graph of balloon diameter vs. temperature. What is the correlation?

Experiment 3: Enzymes in food

This experiment demonstrates the presence of amylase in some common foods. It is important to remember

that amylase breaks down starch and iodine is a starch indicator that turns dark purple in the

presence of starch.

Materials

Starch solution

Iodine‐Potassium iodide (IKI)/

Lugol’s Solution

Empty 2 oz bottle (i.e. corn syrup

bottle)

2 Spray lids

Permanent marker

Paper towel*

Ginger root*

2 or more food products (e.g. apple,

potato)*

*You must provide

Procedure

Note: Test as many foods as you like!

1. Attach the spray lid to the starch solution. The lid will not screw on; simply place it on top.

2. Pour the remaining IKI solution into an empty 2 oz bottle (i.e. corn syrup bottle) and attach a

spray lid. Again, the lid will not screw on, simply place it on top.

Note: When pouring the IKI into the empty bottle, first, be sure the bottle has been thoroughly

cleaned. Also,

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Lab 9: Enzymes

bottle to avoid any possi‐

pour the IKI solution into a graduated cylinder and from there pour it into the empty

ble spills.

3. Spray a paper towel with starch. Let dry for 2 hours.

4. In the mean time, set up a control for this experiment. (Hint: What happens when IKI solution is

mixed with starch? What happens when it is mixed with another liquid?)

5. Cut the food specimens so that a fresh surface is exposed.

6. Gently rub each specimen on the paper towel, back and forth 10‐15 times. Label where each

specimen was rubbed on the paper towel with a permanent marker.

7. Rub a drop of saliva into the paper towel. (We know saliva contains amylase.)

8. Wait 5 minutes.

9. Hold the spray bottle 10‐12 inches from the paper towel, and mist with the IKI solution spray.

10. Observe where color develops, indicating the presence of starch.

Questions

1. What is the function of amylase? What does amylase do to starch?

2. What were your controls for this experiment? What did they prove?

3. Why did you wait 5 minutes before spraying with IKI solution?

4. Which of the foods that you tested contained amylase? Which did not? What experimental

evidence supports your claim?

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Lab 9: Enzymes

5. There is another digestive enzyme other than salivary amylase that is secreted by the salivary

glands. What is this enzyme? What substrate does it act on? Where in the body does

it become activated, and why?

6. Saliva does not contain amylase until babies are about two months old. How could this

affect an infant’s digestion? (Hint: babies do not eat cereal until about three months old)

7. Many common household products contain enzymes (e.g., meat tenderizer, enzyme detergent).

Can you think of any other household uses for enzymes?

8. The stomach contains enzymes that aid in digestion, including proteases which digest proteins.

Why don’t these enzymes digest the stomach and small intestine, which are partially

composed of protein?

103


104


The Cell

Lab 10

Cell Structure & Function

105


106


Lab 10: Cell Structure & Function

Concepts to explore:






What is a cell?

Prokaryotes

Eukaryotes

Cell structure

Function of cell structures

Introduction

A cell is the fundamental unit of life. All living organisms originate from a single cell. Some remain as a

single cell, while others become multi‐cellular (like you!).

Though most cells are difficult to see with the naked eye, using the microscope, cytologists have identified

many of their features. These range from the characteristics of the outer membranes, to internal

structures such as the nucleus and mitochondria and have become the foundation for what is now

known as “cell theory”.

Cell theory states:

All cells are generated from previous cells

All cells pass on their genetic information

All living things are made of cell(s)

Energy metabolism occurs inside cells

The chemical make‐up of cells is similar

Cytologists are scientists who

study cells. The study of the

cell is known as cytology.

Although all organisms are made up of cells, not all cells are identical. Prokaryotes and eukaryotes are

two structurally different types of cells.

Prokaryotes are the most primitive and basic organisms. They lack a membrane

bound nucleus and membrane bound organelles (specialized structures). The term

prokaryote comes from the Latin words “pro” (before) and “karyote” (nucleus).

Eukaryote are much more complex organisms, containing both a nucleus and

membrane bound organelles. The term “eukaryote” comes from the Latin words

“eu” (true) and “karyote” (nucleus). Protists, fungi, plant and animal cells are all

eukaryotic cell(s).

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Lab 10: Cell Structure & Function

Prokaryotes

Cyanobacteria and archaea (both

primitive bacteria) are the only

prokaryotes.

Are very small (.1µm to 2µm)

Reproduce asexually. This

means sexual reproduction is

absent, and there is little genetic

variation between generations

Have simple cellular components

Are capable of living almost anywhere

and often thrive in harsh

conditions

Are unicellular

Eukaryotes

2 billion years younger than prokaryotic

cells

Great biological diversity

All multi‐cellular organisms are

eukaryotes

Significantly larger than most

prokaryotic cells

More complex shapes and internal

structure than prokaryotes

Some are capable of capturing

light energy (chloroplasts in

plant cells and cones and rods of

the eye)

Figure 1: A sample prokaryote

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Lab 10: Cell Structure & Function

NOT SHOWN:

Eukaryotes:

Glycocalyx: A “slime coating” on some prokaryotic cells that helps protect the cell

and enables it to attach to “unconventional” surfaces (i.e. teeth, lungs, artificial

joints).

Thylakoid: Extensions of the plasma membrane of cyanobacteria containing photosynthetic

pigments.

Flagella: Long cylindrical protrusions that rotate to provide mobility.

Pili: Hair like extensions on the cell surface that transmit genetic information and

help secure the bacteria to its host.

Figure 2: Structures unique to eukaryotes

NOT SHOWN:

Nucleolus: A part of the nucleus that is made of RNA, Protein and Chromatin and

manufactures RNA and ribosomes.

Cytoskeleton: The “skeleton” found in all eukaryotic cells that provides shape to

the cell while also enabling it to move. It consists of three parts:

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Lab 10: Cell Structure & Function

Microfilaments: Small strands that help the cell resist tension. Think of it as a

piece of wire.

Intermediate filaments: Anchors the organelles in the cell and provide additional

stability.

Microtubules: Small hollow tubes that help the cell maintain its shape, move

things around within the cell and form other key structures.

Centriole: Barrel shaped structures that help make cilia and flagella. They also play

a key role in cell division.

Cilia: Small “hairs” on the outside of the cell. They help the cell move and are sensory

receptors.

Flagella: The structure of eukaryotic flagella is far more complex than prokaryotic

flagella. They provide mobility by rotating back and forth, they help transport fluids

and serve as sensory receptors.

Mitochondria: The “power plant” of the cell. They are a membrane bound organelle

(inner and outer membrane) with their own circular DNA, and make ATP

(energy) for the rest of the cell.

Chloroplast: Think of them as the plant version of mitochondria. The main difference

is that they take light energy and convert it to mechanical energy.

Peroxisomes: Contain enzymes that help the cell destroy toxins.

Vacuole: Membrane bound “sacs” that provide storage and provide transportation

within the cell (excretion, secretion).

Vesicle: Plays a similar role to vacuoles, but are smaller.

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Lab 10: Cell Structure & Function

Structure Prokaryotic Cell Eukaryotic Cell

Nucleus No Yes

Plasma Membrane Yes Yes

Cell Wall Yes Yes (in most cells)

Cytoplasm Yes Yes

Flagella and Pili

Prokaryotic vs. Eukaryotic Cells

Occasionally

Flagella‐ Occasionally

Pili ‐ No

Cilia No Occasionally

Glycocalyx Occasionally Occasionally

Cytoskeleton No Yes

Endoplasmic Reticulum No Yes

Mitochondria No Yes

Golgi Apparatus No Yes

Chloroplast No In plants and many protists

Ribosome Yes Yes

Lysosome No Yes

Peroxisome No Yes

Vacuole and Vesicle No Yes (in most cells)

Experiment 1: Identifying cell structures

View the slide pictures and images below, paying attention to detail, and note the different characteristics

of prokaryotes and eukaryotes. On each picture, label the parts indicated if they are visible. If

you can not see them, draw and label them where they would be located.

Figure 3

Bacteria: Nucleoid, cell wall, plasma membrane, ribosomes, glycocalyx (if present), flagella (if present)

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Lab 10: Cell Structure & Function

Figure 4

Protist: Nucleus, plasma membrane, cytoplasm, chloroplasts (if present), flagella (if present)

Figure 5

Plant Cell: Nucleus, cell wall, plasma membrane, cytoskeleton, cytoplasm, chloroplast, mitochondria,

vacuoles

Figure 6

Animal Cell: Nucleus, nucleolus, nuclear envelope, plasma membrane, cytoplasm, mitochondria, golgi

apparatus, ER (rough and smooth), ribosome, lysosomes, peroxisomes, vesicles

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Lab 10: Cell Structure & Function

Questions

1. For each structure identified, do you think its location affects its ability to function? Why or

why not? (Hint: those buried deep in the cell probably do different things than those closer

to the cell membrane)

2. Draw a labeled diagram of a small section of the plasma membrane and briefly describe its

structure and function.

3. Describe the differences between animal and plant cells.

4. Which of the following structures are present in both prokaryotic and eukaryotic cells?

Plasma membrane, Golgi apparatus, DNA, lysosomes and peroxisomes, cytoplasm

5. Where is genetic material found in plant cells?

6. Mitochondria contain their own DNA (circular) and have a double membrane. What explanation

for this observation can you come up with?

(Hint 1: Where else do we see circular DNA?)

(Hint 2: What do you know about the relative age of eukaryotic cells?)

7. How is the structure of the cellulose wall related to its function?

8. Defects in structures of the cell can lead to many diseases. Pick one structure of a eukaryotic

cell and develop a hypothesis as to what you think the implications would be if that structure

did not function properly.

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Lab 10: Cell Structure & Function

9. Using books, articles, the internet, etc. conduct research to determine if your hypothesis was

correct.

Experiment 2: Create a cell

In this experiment you will create an animal and a plant cell using household items, to observe the difference

between the two types of cells.

Materials

4 Unflavored gelatin packets

2 Resealable bags

Warm water*

Tupperware*

Bowl*

Household items to use as cell structures*

*You must provide

Procedure

1. Place four packets of unflavored gelatin in a bowl . Prepare according to the directions on

the package, but do not place it in the refrigerator.

2. Locate household items that can serve as the nucleus, mitochondria, ribosomes, ER, golgi

apparatus and chloroplasts.

For example, a plum works great as a nucleus, small mandarin oranges work great as mitochondria,

string works well for the ER, etc. Be creative and come up with your own ideas!

3. Open one resealable bag (these serve as the cell membrane) and pour half of the liquid

gelatin into it.

4. We will first make the plant cell so add the items that you have designated for each cell

structure into the gelatin and tightly close the bag.

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Lab 10: Cell Structure & Function

5. Place the bag in the square disposable Tupperware (this serves as the plant cell wall).

6. Open the other resealable bag and pour the remainder of the gelatin into it.

7. To make the animal cell, add the items you have designated for each cell structure into the

bag and close it tightly.

8. Place both “cells” into the refrigerator for 24 hours.

9. Return after 24 hours and observe the “cells” you have made. Notice the difference between

the animal cell and the plant cell.

Questions

1. What cell structures did you place in the plant cell that you did not place in the animal cell?

2. Is there any difference in the structure of the two cells?

3. What structures do organisms that lack cell walls have for support?

4. How are organelles in a cell like organs in a human body?

5. How does the structure of a cell suggest its function? List three examples.

115


116


The Cell

Lab 11

Mitosis

117


118


Lab 11: Mitosis

Concepts to explore:

Concepts to explore:


Chromosomes


Telophase


Cell cycle


Cytokinesis


Mitosis


Interphase


Metaphase


Anaphase

Introduction

DNA is often referred to as the “code of

life”. Within the nucleus of each cell, it

contains all the information necessary for

If the DNA from a single cell was uncoiled

and stretched out, it would be

over 2 meters (6 feet) long, but too thin

to see.

that cell, or any multicellular organism (you), to function. It is a long, continuous

strand, tightly packed and stored in large molecules called chromosomes

(Figure 1).

Mitosis is the process where somatic cells (non‐sex cells) replicate and divide

their nucleus. Before mitosis occurs (Figure 1), each cell has two complete sets

of chromosomes (one set from your mother and one set from your father –

“2n”). The process of mitosis generates diploid (2n) daughter cells that contain

Figure 1: Human karyotype two complete sets of chromosomes identical to the parent cell. Mitosis is repeated

every time a cell divides,

trillions of times in a human body.

The life of a cell is divided into four stages, which are repeated

until the cell receives instructions to do otherwise.

During this cell cycle, the cells duplicate their genomes,

segregate that information, and divide, producing

daughter cells.

The stages in the cell cycle are:

G 1 : the first growth phase during which the cell

grows and makes the components necessary

for replication

S: DNA replicates

Figure 2: The stages of the cell cycle

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Lab 11: Mitosis

G 2 : the second growth phase in which cellular organelles (mitochondria, ribosomes, and centrioles)

are replicated.

M: mitotic division (prophase, metaphase, anaphase and telophase)

A cell normally completes the cycle in 18‐24 hours, with mitosis occupying 1‐2

hours of that time (Figure 2). Each stage is regulated by specialized proteins

that coordinate the division and cell growth. Certain types of cancer are associated

with the failure of these proteins.

Figure 3: Two chromatids

are joined together by a

centromere to form a

chromosome pair.

Chromosomes are joined as a four‐arm structure by a centromere (Figure 3).

During cell division (both meiosis and mitosis) each chromosome is duplicated,

resulting in two identical chromatids. For a short while the cell contains four

copies of each chromosome (two from your mother and two from your father).

The chromatids are then pulled apart and divided into daughter cells.

In mitosis, the division of the parent cell produces two cells, each having two sets of chromosomes

(diploid (2n)). Thus, somatic cells can replicate and maintain the right number of chromosomes.

Mitosis (Figure 4):






Interphase: The longest period of the cell cycle was named Interphase because it was the

“in‐between” period between cell cycles. It is the phase where the cell grows, its DNA replicates

and it prepares for division. The replicated chromosomes pair up as sister chromatids

(exact copies of each other).

Prophase: The nuclear membrane breaks down and the chromosomes separate. Structures

which will serve as anchors in the cell (centrioles) during the division process appear.

Metaphase: The chromosomes line up in the middle of the cell. Microtubules attach to the

chromosomes. The orientation of each pair of homologous chromosomes is independent

from all other chromosomes. This means they can “flip flop” as they line up, effectively

shuffling their genetic information into new combinations. Microtubules (long strands)

grow from each centriole and link together while also attaching to each pair of homologous

chromosomes.

Anaphase: The microtubules pull the sister chromatids apart.

Telophase: One set of chromosomes arrives at each centriole, at which time a nucleus

forms around each set.

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Lab 11: Mitosis

Cytokinesis: The plasma membrane of the cell folds in and encloses each

nucleus into two new diploid daughter cells.

Figure 4: The stages of mitosis

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Lab 11: Mitosis

Experiment 1: Observation of mitosis in a plant cell

Mitosis is virtually identical in plant and animal cells, however, there is one small difference which occurs

during telophase. Plants, due to the presence of a cell wall, can not “pinch” the cytoplasm into

two daughter cells. Instead, a new cell wall must be developed which will then separate the two cells,

allowing them both to be fully covered with a cell wall.

In this experiment we will look at the different stage of mitosis in an onion cell. The large size of the

onion cell allows the different stages of mitosis to be observed with the aid of a microscope. Also,

when you are asked in the lab to specify the amount of time each stage takes during the cell cycle, remember

that mitosis only occupies 1‐2 hours while interphase can take anywhere from 18‐24. With

this information, you will be asked in the procedure below to calculate the percentage of cells in each

stage of the cell cycle.

Materials

Prepared Allium root tip digital slide picture (Figure 5)

Procedure

1. The length of the cell cycle in the onion root tip is about 24 hours. Predict how many hours

of the 24 hour cell cycle you think each step takes in Table 1.

2. Examine a digital slide picture (following page) of an onion root tip. Count the number of

cells in each stage within a single field of view. Record this number in Table 1.

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Lab 11: Mitosis

Figure 5: Onion root tip

slide

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Lab 11: Mitosis

Table 1: Number of cells observed in each stage of the cell cycle

Stage Predicted %

Interphase

Number of cells in

stage

Total number of cells in

field

Calculated %

Prophase

Metaphase

Anaphase

Telophase

Totals

3. Reexamine the onion root tip slide at low magnification. Locate the region just above the root

cap (use digital slide picture on previous page if you do not have the slide set).

4. Focus on this zone of the onion root tip and locate the stages of the cell cycle.

5. Using the space below, draw the dividing cell in the appropriate area for each stage of the

cell cycle.

Mitosis

Interphase

Prophase

Prometaphase

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Lab 11: Mitosis

Metaphase

Anaphase

Telophase

Questions

1. What stage were most of the onion root tip cells in? Does this make sense?

2. As a cell grows, what happens to its surface area : volume ratio? (Think of a balloon being

blown up). How is this changing ration related to cell division?

3. What is the function of mitosis in a cell that is about to divide?

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Lab 11: Mitosis

4. How accurate were your time predictions for each stage of the cell cycle?

5. Discuss one observation that you found interesting while looking at the onion root tip cells.

6. What would happen if mitosis were uncontrolled?

126


The Cell

Lab 12

Meiosis

127


128


Lab 12: Meiosis

Concepts to explore:

Concepts to explore:





Meiosis

Diploid cells

Haploid cells

Chromosomal crossover

Introduction

Meiosis only occurs in organisms that reproduce sexually. The process generates haploid (1n) cells

called gametes (sperm cells in males and egg cells in females),

or spores in some plants, fungi, and protists, that

contain one complete set of chromosomes. Haploid cells

fuse together during fertilization to form a diploid cell with

two copies of each chromosome (2n).

There are over two meters of DNA packaged

into a cell’s nucleus. It is coiled and

folded into superhelices that form chromosomes,

which must be duplicated before

a cell divides.

Genes are the units of heredity that have specific loci

(locations) on the DNA strand and code for inheritable

traits (such as hair color). Alleles are alternative forms of the same gene (brown vs. blue eyes). Homologous

chromosomes contain the same genes as each other but often different alleles. Non‐sex cells

(e.g. bone, heart, skin, liver) contain two alleles (2n), one from the sperm and the other from the egg.

Mitosis and meiosis are similar in many ways. Meiosis, however, has two rounds of division—meiosis I

and meiosis II. There is no replication of the DNA between meiosis I and II. Thus in meiosis, the parent

cell produces four daughter cells, each with just a single set of chromosomes (1n).

Meiosis I is the reduction division– the homologous pairs of chromosomes are separated so that each

daughter cell will receive just one set of chromosomes. During meiosis II, sister chromatids are separated

(as in mitosis).

Each of the 23 human chromosomes

has two copies. For each chromosome,

there is a 50:50 chance as to which copy

each gamete receives.

That translates to over 8 million possible

combinations!

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Lab 12: Meiosis

Meiosis:











Prophase I: The sister chromatids attach to their homologous counterparts (same chromosome

– different version). This is the stage where crossing over occurs (homologous chromosomes

exchange regions of DNA). Structures which will serve as anchors in the cell

(centrioles) during the division process appear.

Metaphase I: The chromosomes line up in the middle of the cell. The orientation of each

pair of homologous chromosomes is independent from all other chromosomes. This

means they can “flip flop” as they line up, effectively shuffling their genetic information

into new combinations. Microtubules (long strands) grow from each centriole and link

them together while also attaching to each pair of homologous chromosomes.

Anaphase I: The microtubules pull the homologous chromosomes apart (the sister chromatids

remain paired).

Telophase I: One set of paired chromosomes arrives at each centriole, at which time a nucleus

forms around each set.

Cytokinesis: The plasma membrane of the cell folds in and encloses each nucleus into two

new daughter cells.

Prophase II: Before any replication of the chromosomes can take place, the daughter cells

immediately enter into prophase II. New spindle fibers form as the nucleus breaks down.

Metaphase II: The sister chromatids align in the center of the cell, while the microtubules

join the centrioles and attach to the chromosomes. Unlike metaphase I, since each pair of

sister chromatids is identical, their orientation as they align does not matter.

Anaphase II: The sister chromatids are separated as the microtubules pull them apart.

Telophase II: The chromatids arrive at each pole, at which time a nucleus forms around

each.

Cytokinesis: The plasma membrane of the cell folds in and engulfs each nucleus into two

new haploid daughter cells.

We briefly discussed “crossing over” in Prophase I. Since the chromosomes of each parent undergoes

genetic recombination, each gamete (and thus each zygote) acquires a unique genetic fingerprint.

The closeness of the chromatids during prophase I, creates the opportunity to exchange genetic material

(chromosomal crossover) at a site called the chiasma. The chromatids trade alleles for all genes

located on the arm that has crossed.

The process of meiosis is complex and highly regulated. There are a series of checkpoints that a cell

must pass before the next phase of meiosis will begin. This ensures any mutated cells are identified

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Lab 12: Meiosis

and repaired before the cell division process can continue.

One of the mutations that is of particular concern is a

variation in the amount of genetic material in a cell. It is

critical that the gamete contain only half of the chromosomes

of the parent cell. Otherwise the amount of DNA

would double with each new generation. This is the key

feature of meiosis.

Mutations that are not caught by the cell’s

self‐check system can result in chromosomal

abnormalities like Down’s syndrome, in

which there are 3 copies of chromosome

21.

Figure 1: The stages of meiosis

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Lab 12: Meiosis

Experiment 1: Following chromosomal DNA movement

Every cell in the human body has two alleles that condense into single chromosomes held together by

a centromere. These “sister” chromatids replicate and pair with the newly made homologous chromosomes.

In this exercise we will follow the movement of the chromosomes through meiosis I and II to

create haploid (gamete) cells.

Materials

2 sets of different colored snap

beads (32 of each)

8 centromeres (snap beads)

Blue and red markers*

*You must provide

Figure 2: Bead Set‐up

Procedure

Meiosis I

A. As prophase I begins, chromosomes coil and condense in preparation for replication.

1. Using one single color of bead, build a homologous pair of duplicated chromosomes.

Each chromosome will have 10 beads with a different colored centromere in it.

For example, if there are 20 red beads, 10 beads would be snapped together to

make two different strands. In the middle of each of the 10 bead strands, snap

a different colored bead in to act as the centromere.

Now, repeat these steps using the other color of bead.

2. Assemble another homologous pair of chromosomes using only 12 (that’s 6 per

strand) of the first color bead. Place another, different colored bead in the middle of

each to act is its centromere. Repeat this step (2 strands of 6 beads plus a centro‐

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Lab 12: Meiosis

mere) with the other color of beads.

B. Bring the centromeres of two units of the same color and length together so they can be held

together to appear as a duplicated chromosome.

1. Simulate crossing over. Bring the two homologues pairs together (that’d be the two

pairs that both have 10 bead strands) and exchange an equal number of beads between

the two.

C. Configure the chromosomes as they would appear in each of the stages of meiosis I.

Meiosis II

A. Configure the chromosomes as they would appear in each stage of meiosis II.

B. Return your beads to their original starting position and simulate crossing over. Track how this

changes the ultimate outcome as you then go through the stages of meiosis I and II.

C. Using the space below, and using blue and red markers, draw a diagram of your beads in each

stage. Beside your picture, write the number of chromosomes present in each cell.

Meiosis I

Prophase I

Metaphase I

Anaphase I

Telophase I

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Lab 12: Meiosis

Meiosis II

Prophase II

Metaphase II

Anaphase II

Telophase II

Questions

1. What is the state of the DNA at the end of meiosis I? What about at the end of meiosis II?

2. Why are chromosomes important?

3. How are Meiosis I and Meiosis II different?

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Lab 12: Meiosis

4. Name two ways meiosis contributes to genetic recombination.

5. Why do you use non‐sister chromatids to demonstrate crossing over?

6. How many chromosomes were present when meiosis I started?

7. Why is it necessary to reduce the chromosome number of gametes, but not other cells of an

organism?

8. If humans have 46 chromosomes in each of their body cells, determine how many chromosomes

you would expect to find in the following:

Sperm ___________________

Egg ___________________

Daughter cell from mitosis ___________________

Daughter cell from Meiosis II ___________________

9. Investigate a disease that is caused by chromosomal mutations. When does the mutation

occur? What chromosome is affected? What are the consequences?

135


136


The Cell

Lab 13

DNA & RNA

137


138


Lab 13: DNA & RNA

Concepts to explore:

Concepts to explore:







DNA structure

Nucleotides

Amino acids

Proteins

Genetic code

Mutation




RNA

Transcription

Translation

Introduction

Long before we had any understanding of how, we knew that traits were

passed on from generation to generation. We knew traits were expressed

as heritable proteins, but we had no idea of the mechanism.

Whatever the mechanism, it needed to meet three criteria:

It needed to carry information between generations.

It needed to express that information.

It needed to be easily replicable.

Prior to the 1950’s, there was much debate over what the structure of a

molecule that met all three criteria would look like. Though a number of

people made significant contributions, in 1953 James Watson and Francis

Crick won the Nobel Prize for their model of what we now know as DNA

(deoxyribonucleic acid). The features of this model satisfied all of the

necessary criteria.

Figure 1: DNA Double Helix

DNA takes the form of what is commonly referred to as a “double helix”, or perhaps more simply, a long

twisted ladder with rungs (Figure 1). The sides of the ladder consist of a sugar‐phosphate “backbone”

and the strand itself has directionality. In other words, like the words on this page, there is a set order

in which they are read. In the case of DNA, it is from the 5’ (five

prime) to 3’ (three prime) end.

Figure 2: Nucleotides

The rungs of the ladder carry information in a sequential series of four

different nucleotides (small molecules): Guanine (G), Adenine (A),

Thymine (T) and Cytosine (C). These nucleotides pair up in a very precise

manner (specificity); A with T, and G with C (Figure 2). No other

combinations are ever made because of the chemical and electrical

forces within the nucleotide.

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Lab 13: DNA & RNA

As a cell divides, the DNA double helix splits

into a single helix (Figure 3). Each single helix

then serves as a template for a new strand.

Neighboring nucleotides then bind to the single

strand helix after which a new sugarphosphate

backbone is formed.

The specificity in which the nucleotides pair

means the two new double helices (DNA) are

identical to the original. It is the sequence of

these nucleotides that are passed on from one

generation to another, as heritable information.

Figure 3: DNA replication

So the question remains, why is the sequence of these different four nucleotides so important? Simply

put, they instruct your cells what proteins to make and how to make them (your body is made of proteins).

If the protein is wrong you are likely either very sick or dead.

Proteins are simply chains of amino acids (small molecular building blocks) that are linked together.

Twenty different amino acids are available to produce all the proteins in the body. Each amino acid is

coded for by a three nucleotide sequence (codon). The sequence of the amino acids determines the

size of the protein and how it will fold, both factors that determine its function. Other factors such as

charge and hydrophobicity (an aversion to water molecules) play a role in determining how a protein

folds. Consider the following analogy:

“The earth revolves around the sun.”

Each of the 12 different letters (codons) in the preceding sentence is largely uninformative.

When letters are assembled they create words, which have meaning.

Linked words create a sentence (protein), which is then informative.

Consider a protein that is five amino acids long. Picking from the 20 available amino acids there are 5 20

different possible combinations (3,200,000). Even small proteins are typically several hundred amino

acids long. The number of different proteins that can ultimately be coded for by 20 amino acids is virtually

limitless.

The next obvious question is how do the 4 nucleotides “code” for 20 different amino acids. Each

“letter” (codon) in the genetic code is made up of three nucleotides which codes for a specific amino

acid. If we start with four possible nucleotides (A, T, G, C), how does your body make twenty different

amino acids?

If the “letter” is two nucleotides long, there are 16 possible “letters” (2 4 ) ‐ not enough.

If the “letter” is three nucleotides long, there are 64 possible combinations (3 4 ) ‐ more than

what’s needed for twenty amino acids.

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Lab 13: DNA & RNA

Like a sentence, the reader (a cell) needs to know where to start and where to stop (two more codons,

for a total of 22). The remaining 42 possible combinations make up what is referred to as “the redundancy

of the code”. In other words; Tim, Tom, Tam would all be the same person, it is simply three different

spellings for his name. Each combination of three nucleotide is known as a “codon”.

Experiment 1: Coding

Materials

Red beads

Blue beads

Yellow beads

Green beads

For the following exercises:



Regular beads are used as

nucleotides

Pop‐it beads are used as

amino acids

Procedure

A) Using red, blue, yellow and green beads, devise and lay out a three color code for each

of the following letters (codon). For example Z = green:red:green.

In the spaces below the letter, record your “code”.

C: E: H: I: K: L:

_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _

M: O: S: T: U:

_ _ _ _ _ _ _ _ _ _ _ _ _ _ _

Create codons for: Start: Stop: Space:

_ _ _ _ _ _ _ _ _

B) Using this code, align the beads corresponding to the appropriate letter to write the following

sentence (don’t forget start, space and stop):

The mouse likes most cheese

1. How many beads did you use?

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Lab 13: DNA & RNA

There are multiple ways your cells can read a sequence of DNA and build slightly different proteins from

the same strand. We will not go through the process here, but as an illustration of this “alternate splicing”,

remove codons (beads) 52‐66 from your sentence above.

2. What does the sentence say now? (re‐read the entire sentence)

Mutations are simply changes in the sequence of nucleotides. There are three ways this occurs:

Change one, remove one, or add one

Using the sentence from exercise 1B:

C) Change the 24 th bead to a different color.

3. What does the sentence say now? (re‐read the entire sentence)

4. Does it make sense?

D) Replace the 24 th bead and remove the 20 th bead (remember what was there).

5. What does the sentence say (re‐read the entire sentence)?

6. Does it make sense?

7. Where does it make sense?

E) Replace the 20 th bead and add one between bead numbers 50 and 51.

8. What does the sentence say?

9. Does it make sense?

10. In “C” we mutated one letter. What role do you think the redundancy

of the genetic code plays, in light of this change?

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Lab 13: DNA & RNA

11. Based on your observations, why do you suppose the mutations we made

in “D” and “E” are called frame shift mutations.

12. Which mutations do you suspect have the greatest consequence? Why?

DNA codes for all of the proteins manufactured by any organism (including you!). It is valuable, highly

informative and securely protected in the nucleus of every cell.

Consider the following analogy:

An architect spends months or years designing a building. His original drawings are valuable and

informative. He will not provide the original copy to everyone involved in constructing the

building.

Instead, he gives the electrician a copy with the information he needs to build the electrical system.

He will do the same for the plumbers, the framers, the roofers and everyone else who

needs to play a role to build the structure. These are subsets of the information contained in

the original copy. Your cell does the same thing. The “original drawings” are contained in your

DNA which is securely stored in the nucleus.

Nuclear DNA is “opened up” by an enzyme (Helicase) and a subset of information is transcribed (copied)

into RNA. RNA is a single strand version of DNA, where the nucleotide uracil, replaces thymine. The copies

are sent from the nucleus to the cytoplasm in the form of mRNA (messenger RNA). Once in the cytoplasm,

tRNA (transfer RNA) links to the codons and aligns the proper amino acids, based on the mRNA

sequence.

The ribosomes (protein builders) that float around in the cytoplasm, latch onto the strand of mRNA and

sequentially link the amino acids together that the tRNA has lined up for them. This construction of proteins

from the mRNA is known as translation.

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Lab 13: DNA & RNA

Experiment 2: Transcription and translation

Materials

Red beads

Blue beads

Green beads

Yellow beads

Pop‐it beads (8 different colors)

Procedure

A) Write a five word sentence using no more than 8 different letters.

B) Using the four colored beads, create “codons” (3 beads) for each letter in your sentence,

plus ones for “start, “space” and stop”.

C) “Write” the sentence using the beads.

1. How many beads did you use?

Using your pop‐it beads, assign one bead for each codon (you do not need beads for start, stop

and space). These will be your amino acids.

Connect the “Pop it” beads to build the chain of amino acids that codes for your sentence

(leave out the “start”, “stop” and “space”).

2. How many different amino acids did you use?

3. How many total amino acids did you use?

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Lab 13: DNA & RNA

Experiment 3: DNA Extraction

Materials

Fresh soft fruit (i.e. grapes, berries

or banana)*

Cheesecloth

Rubber band

Plastic zipper bag

2 100mL Beaker

DNA extraction solution**

Standing test tube

Scissors*

Ethanol (ice cold)***

Stir stick

Rubber band

*You must provide

** Sodium chloride, detergent and water

***For ice cold ethanol, store in the freezer

60 minutes before use.

Figure 4: DNA extracted from fruit was dyed

with a substance that glows under black light.

Procedure

1. Put pieces of soft fruit (approximate size of 5 grapes) into a plastic zipper bag and mash

with your fist.

2. Using a 100ml beaker, measure out 10ml of the DNA extraction solution and pour it into

the bag with the fruit it in. Seal the bag completely.

3. Mix well by kneading the bag for 2 minutes.

4. Create a filter by placing the center of the cheesecloth over the mouth of the standing test

tube, pushing it into the tube about 2 inches, and securing the cheesecloth with a rubber

band around the top of the test tube.

5. Cut a hole in the corner of the bag and filter your extraction by pouring it into the cheese‐

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Lab 13: DNA & RNA

cloth (the filtered solution in the beaker is what you keep).

6. While holding the test tube at a 45° angle, slowly pour 5ml ice‐cold ethanol into the test

tube.

7. As the DNA enters the ethanol it will precipitate (come out of solution). Let the test tube sit

for 2‐3 minutes. You should see air bubbles on DNA, which will eventually float to the top

of the ethanol.

8. Gently insert the stir stick into the test tube and slowly raise and lower the tip several

times, to spool and collect the DNA.

Questions

1. Which DNA bases pair with each other?

2. How is information to make proteins passed on through generations?

3. Why did we use a salt in the extraction solution?

4. What else might be in the ethanol/aqueous interface? How could you eliminate this?

5. What is the texture and consistency of DNA?

6. Is the DNA soluble in the aqueous solution or alcohol?

7. What surprised you about DNA replication and protein synthesis?

146


The Cell

Lab 14

Mendelian Genetics

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Lab 14: Mendelian Genetics

Concepts to explore:

Gregor Mendel

Law of segregation

Homozygous

Heterozygous

Independent assortment

Dominant vs. recessive








Incomplete dominance

Co‐dominance

Genotype

Phenotype

Monohybrid cross

Dihybrid cross

Punnett square

Introduction

In 1866, Gregor Mendel, an Austrian Monk, published a paper entitled “Experiments in plant hybridization”.

It went largely unnoticed until 1900 when it was rediscovered and subsequently became the

basis for what we now refer to as Mendelian Genetics.

Mendel was the first to recognize:



Inherited characters are determined by specific factors (now recognized these as genes).

These factors occur in pairs (genes).

When both alleles of a gene are the same they are said to be homozygous, while if they are different

they are said to be heterozygous. When gametes form, these factors segregate so that each gamete

contains only one allele for each gene. Remember, alleles reside on the chromosomes that are dividing.

These original observations lead to what we now refer to as The law of segregation and the law of

independent assortment.

Figure 1: Law of Segregation

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Lab 14: Mendelian Genetics

The law of segregation states that during

meiosis, homologous (paired) chromosomes

split (Figure 1). The law of independent

assortment states that during

meiosis, each homologous chromosome

has an equal chance of ending up in either

gamete, and alleles for individual

genes segregate with the chromosomes

on which they are located (Figure 2).

Using corn as an example (Figure 2):

Figure 2: Law of Independent Assortment



The large chromosome has the gene for kernel color (Y = yellow, y = blue).

The small chromosome has the gene for kernel texture (S = smooth (green); s = wrinkled (red)).

When a dominant allele is present, that characteristic is expressed, regardless of the second allele. In

this case both the Yy and YY offspring will be yellow.

A recessive allele is only expressed when both alleles are recessive. In this case only the yy combination

is blue. The dominant allele is always represented by capital letters, while the recessive is represented

by lower case letters.

Genotype refers to the combination of alleles for a particular trait. Phenotype refers to the appearance

of that combination of alleles. In our example, the genotype of the diploid cell is Yy, Ss, while the

phenotype is Yellow and Smooth.

Figure 3: Monohybrid Cross

Punnett Square F 1

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Lab 14: Mendelian Genetics

Parent 2

Parent 1

Y

Y

y Y y Y y

Parent 2

Y

Parent 1

Y Y Y Y y

y

y Y y Y y

y Y y y y

Figure 4: Punnet square of a monohybrid cross

(F 1 )

Figure 5: Punnet square of a monohybrid cross

(F 2 )

Y s Y S y S y s

Y s Y Y s s Y Y S s y Y S s y Y s s

Y S Y Y s S Y Y S S y Y S S y Y s S

y S Y y s S Y y SS y y S S y y s S

y s Y y s s Y y S s y y S s y y s s

Figure 6: Punnet square of a dihybrid cross (F 1 )

Alleles can exhibit incomplete dominance and co‐dominance. An example of incomplete dominance is

the cross of two plants, one with red flowers and one with white, whose offspring have pink flowers.

In the case of codominance, the same cross would result in red and white striped flowers.

If we know the genotype of two parents we can predict both the genotype and phenotype of their offspring

using a Punnett Square. A monohybrid cross is a cross between two parents (P), looking at a

single gene (Figure 3). In this example, both parents are pure breeding (homozygous); one for the yellow

color and one for the blue color. This cross can be shown as a Punnett Square (Figure 4), with each

parent (P) contributing a single gamete. The offspring (F 1 ) are determined adding the gamete of each

parent (P) (Row and Column). The F 1 genotypes are all Y , y; with yellow phenotypes. The cross of the

(F 1 ) generation, known as the F 2 generation, is shown in Figure 5. The Punnett square can also predict

the F 1 for multiple genes.

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Lab 14: Mendelian Genetics

Using our corn example, let’s look at two genes (color and texture), also known as a dihybrid cross. In

this example we use two parents that are heterozygous for both traits (Figure 6), using the gametes we

already identified in (Figure 2).

The F 2 phenotypes are:

Yellow & Smooth: 9

Yellow & Wrinkled: 3

Blue & Smooth: 3

Blue & Wrinkled: 1

NOTE:

9: 3: 3: 1

Is a ratio you should

remember for question

#4

Experiment 1: Punnett square crosses

Materials

Red beads

Blue beads

Green beads

Yellow beads

2 100mL Beakers

Procedure

1. Set up and complete Punnett squares for each of the following crosses: (remember Y = yellow,

and y = blue)

Y Y and Y y

Y Y and y y

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Lab 14: Mendelian Genetics

a) What are the resulting phenotypes?

b) Are there any blue kernels? How can you tell?

2. Set up and complete a Punnett squares for a cross of two of the F 1 from 1b above:

a) What are the genotypes of the F 2 generation?

b) What are their phenotypes?

c) Are there more or less blue kernels than in the F 1 generation?

3. Identify the four possible gametes produced by the following individuals:

a) YY Ss: ______ ______ ______ ______

b) Yy Ss: ______ ______ ______ ______

c) Create a Punnett square using these gametes as P and determine the genotypes of the

F 1 :

d) What are the phenotypes? What is the ratio of those phenotypes?

4. You have been provided with 4 bags of different colored beads. Pour 50 of the blue beads and

yellow beads into beaker #1 and mix them around. Pour 50 of the red beads and green beads

in beaker #2 and mix them.

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Lab 14: Mendelian Genetics

Attention! Do not pour the beakers together.





#1 contains beads that are either yellow or blue.

#2 contains beads that are either green or red.

Both contain approximately the same number of each colored bead.

These colors correspond to the following traits (remember that Y/y is for kernel

color and S/s is for smooth/wrinkled):

Yellow (Y) vs. Blue (y) Green (S) vs. Red (s).

A. Monohybrid Cross: Randomly (without looking) take 2 beads out of #1.




This is the genotype of individual #1, record this information. Do not put these

beads back into the beaker.

Repeat this for individual #2. These two genotypes are your parents for the next

generation. Set up a Punnett square and determine the genotypes and phenotypes

for this cross.

Repeat this process 4 times (5 total). Put the beads back in their respective beakers

when finished.

a) How much genotypic variation do you find in the randomly picked parents of your

crosses?

b) How much in the offspring?

c) How much phenotypic variation?

d) Is the ratio of observed phenotypes the same as the ratio of predicted phenotypes?

Why or why not?

e) Pool all of the offspring from your 5 replicates. How much phenotypic variation do

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Lab 14: Mendelian Genetics

you find?

f) What is the difference between genes and alleles?

g) How might protein synthesis execute differently if there a mutation occurs?

h) Organisms heterozygous for a recessive trait are often called carriers of that trait.

What does that mean?

i) In peas, green pods (G) are dominant over yellow pods. If a homozygous dominant

plant is crossed with a homozygous recessive plant, what will be the phenotype of

the F 1 generation? If two plants from the F 1 generation are crossed, what will the

phenotype of their offspring be?

B. Dihybrid Cross: Randomly (without looking) take 2 beads out of beaker #1 AND 2 beads out of

beaker #2.

These four beads represent the genotype of individual #1, record this information.

Repeat this process to obtain the genotype of individual #2.

a) What are their phenotypes?

b) What is the genotype of the gametes they can produce?


Set up a Punnett square and determine the genotypes and phenotypes for this

cross.

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Lab 14: Mendelian Genetics

c) What is your predicted ratio of genotypes? Hint: think back to our example dihybrid

cross


Repeat this process 4 times (for a total of 5 trials).

d) How similar are the observed phenotypes in each replicate?

e) How similar are they if you pool your data from each of the 5 replicates?

f) Is it closer or further from your prediction?

g) Did the results from the monohybrid or dihybrid cross most closely match your

predicted ratio of phenotypes?

h) Based on these results; what would you expect if you were looking at a cross of

5, 10, 20 independently sorted genes?

i) Why is it so expensive to produce a hybrid plant seed?

j) In certain bacteria, an oval shape (S) is dominant over round and thick cell

walls (T) are dominant over thin. Show a cross between a heterozygous oval,

thick cell walled bacteria with a round, thin cell walled bacteria. What are the

phenotype of the F 1 and F 2 offspring?

5. The law of independent assortment allows for genetic recombination. The following equation

can be used to determine the total number of possible genotype combinations for any particular

number of genes:

2 g = Number of possible genotype combinations (where “g” is the number of genes)

1 gene: 2 1 = 2 genotypes

2 genes: 2 2 = 4 genotypes

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Lab 14: Mendelian Genetics

3 genes: 2 3 = 8 genotypes

Consider the following genotype:

Yy Ss Tt

We have now added the gene for height: Tall (T) or Short (t).

a) How many different gamete combinations can be produced?

b) Many traits (phenotypes), like eye color, are controlled by multiple genes. If

eye color were controlled by the number of genes indicated below, how many

possible genotype combinations would there be?

5:

10:

20:

157


158


The Cell

Lab 15

Population Genetics

159


160


Lab 15: Population Genetics

Concepts to explore:

Concepts to explore:


Gene pool


Mutation


Gene frequency


Natural selection


Genetic variation


Genetic drift


Founder effect

Introduction

In the previous lab we looked at how genes are passed on to

offspring. In this lab, the exercises are designed to look at individual

genes (two alleles, one dominant, one recessive). However,

we will be looking at their presence, prevalence and distribution

at the population level.

The gene pool is the sum of all genes and their corresponding

alleles in a given population.

Take a look at the population of 100 brown and white mice in

Figure 1. The color brown (B) is dominant. The standard for

naming alleles is to use the case of the dominant trait, with the

lower case to represent the recessive allele. Their gene pool is

B, b.

Figure 1: Mouse Population

Gene frequency refers to how many times each allele is found

in the population. These 100 mice have 200 genes:


55 heterozygous mice (B, b) have 55 B alleles and

55 b alleles.

27 homozygous recessive mice (b, b) have 54 b alleles (2 x 27 “b”= 54).

18 homozygous dominant mice (B, B) have 36 B alleles (18 x 2 “B” = 36).

The gene frequency of the population is:

B: 91 b: 109

Often this is represented as a percentage of the dominant gene, in this case, the percentage of B is

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Lab 15: Population Genetics

45.5% (=91/200). Note that the dominant gene is less prevalent

than the recessive gene. This is not unusual, it is important to

remember that dominance has no direct relationship to prevalence.

Questions

1. What is the gene pool of this population?

2. What is the gene frequency?

Genetic variation is simply the genetic difference within or between populations, in the gene pool and/

or gene frequency.

Consider the following two populations of butterflies:

Fact: Both populations contain the same 4 colors of butterflies,

thus the gene pool is the same. However, the distribution of

colors within that population is different, thus their gene frequencies

are different.

NOTE: In these exercises on gene

pool, gene frequency and genetic

diversity; assume there are 4 alleles

for color and that all butterflies are

homozygous.

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Lab 15: Population Genetics

Experiment 1: Genetic variation

Materials

Blue beads

Red beads

Green beads

Yellow beads

2 100mL Beakers

2 250mL Beakers

NOTE: When done return beads

to their respective beakers (1 or

2).

Procedure

1. Pour 50 blue beads and 50 red beads into a 250 ml beaker. Without looking, randomly take

50 beads from the 250 ml beaker and place them in a 100 ml beaker (this is beaker #1).

2. Pour 50 green beads and 50 yellow beads into a second 250 ml beaker. Without looking,

randomly take 20 beads from the 250 ml beaker and place them in the other 100 ml beaker

(this is beaker #2).

Questions

1. What is the gene pool of beaker #1?

2. What is the gene pool of beaker #2?

3. What is the gene frequency of beaker #1?

4. What is the gene frequency of beaker #2?

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Lab 15: Population Genetics

5. What can you say about the genetic variation between these populations?

Genetic drift, the variation of the gene pool and/or gene frequency of a population, can result from a

variety of stochastic (random) means. Consider the following population of butterflies who have half

of their habitat destroyed by wildfire:

The remaining population has 50% of the initial gene pool (2 colors) and the gene frequency is different.

As these individuals reproduce, their offspring will no longer reflect the original population.

Experiment 2: Genetic drift

Materials

Blue Beads

Red Beads

Green Beads

Yellow Beads

Beads in beaker #1

Beads in beaker #2

1 100 mL beaker

1 250 mL beaker

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Lab 15: Population Genetics

Procedure

1. From the 250 ml beaker containing green and yellow beads, take 10 beads and place them into

the unused 100 ml beaker (this is beaker #3).

2. Remove half of the beads from beakers #1, #2, and #3 (keep them separated so they can be returned

to the proper beaker). These are the individuals that survived the fire.

3. Record your results and place the beads back in their respective beakers.

4. Repeat this process 4 more times (5 total).

Questions

1. What observations can you make regarding the gene pool and gene frequency of the surviving individuals?

2. What determines how often a phenotype occurs in a population?

3. Are dominant characteristics more frequent in a population than recessive characteristics? Why or

why not?

4. If a selection pressure was against the trait of the dominant allele, what change in a population

would you expect to see?

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Lab 15: Population Genetics

Experiment 3: Founder effect

The same population of butterflies is in the path of a hurricane. All survive, but 10 are blown to a new

location. These 10 start a new population, their progeny will reflect the founders gene pool. This is

known as the founder effect.

Materials

Beaker #1

Beaker #2

Beaker #3

Note: When you are finished with

this experiment, return the beads

to their appropriate bags.

Procedure

1. Remove 10 individuals from beaker #1, 5 from beaker #2 and 2 from beaker #3. These are the

founders of your new population.

2. Record your results and place the beads back in their respective beakers.

3. Repeat this process 4 more times (for a total of 5 trials).

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Lab 15: Population Genetics

Questions

1. What observations can you make regarding the gene pool and gene frequency of the founding

individuals?

2. Do these results vary between the populations founded by beakers #1, #2 and #3? Why or

why not?

3. What observations can you make about the genetic variation between the parent and founding

populations?

Experiment 4: Mutations

Many stochastic events change both the gene pool and gene frequencies over time. These parameters

can also change as a result of mutation and natural selection. Mutations are a change in the sequence

of DNA. Most mutations do not change the phenotype and confer no advantages or disadvantages to

the individual. Each of us has hundreds and probably thousands of mutations that do not affect our

fitness.

To answer the assessments for this exercise, assume the following:




There are approximately 3,000,000,000 base pairs in the mammalian genome (genes constitute

only a small portion of this total).

There are approximately 10,000 genes in the mammalian genome.

A single gene averages 10,000 base pairs in size.

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Lab 15: Population Genetics

Questions

1. How many total base pairs are in all the mammalian genes?

2. What proportion (%) of the total genome does this represent?

3. What is the probability that a random mutation will occur in any given gene?

4. Only 1 out of 3 mutations that occur in a gene result in a change to the protein structure. What

is the probability that a random mutation will change the structure of a protein?

Some mutations do change the protein coded for by a gene. The vast majority of these mutations are

lethal and the embryo never fully develops. Occasionally mutations do not effect embryonic development

and the offspring is born without complication.

Natural selection is a selection pressure that acts on phenotypes in one of three ways:



It will confer an adaptive advantage, an adaptive disadvantage, or remain entirely neutral.

A classic example to illustrate natural selection comes from England.

Prior to the Industrial Revolution the native moths were predominantly a light color, though

darker versions of the same species existed.

The lighter color blended with the light bark of the local trees, while the darker moths experienced a

higher predation rate – they were easier for birds to spot and fewer survived to reproduce. As England

entered the Industrial Revolution they began burning fossil fuels with little regard to the pollutants

they were emitting. The trunks of the trees became coated with soot and the color darkened. The

lighter moths became more conspicuous and the darker were better camouflaged. The proportion of

white to dark moths changed.

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Lab 15: Population Genetics

Experiment 5: Natural selection

Materials

Red beads

Blue beads

1 100mL Beaker

Note: When you are finished with this

experiment, return the beads to their

appropriate bags.

Procedure

1. Remove the two sheets of paper with blue and red “habitats”, at the end of this lab.

2. Place 50 red and 50 blue beads into a 100 ml beaker.

3. Mix them well and pour them onto the sheet marked “Red Habitat”.

4. Keep the beads that fall onto habitat that matches their color.

5. For each bead that you keep (and return to the beaker), add another bead of the same color to

the beaker (discard the rest).

6. Repeat this three times.

7. Record the remaining colors.

Blue _____________

Red _____________

8. With the remaining beads, repeat the process using the “Blue Habitat”.

What beads remain?

Blue _____________

Red _____________

Questions

1. Do you observe a selective advantage or disadvantage for the red or blue beads on the blue

habitat? Why?

2. How did the distribution of phenotypes change over time?

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Lab 15: Population Genetics

3. What results would you predict if starting with the following population sizes?

1000

100

10

Experiment 6: Sickle cell anemia inheritance patterns

Facts:






Sickle Cell Anemia is a genetic disease (1 base pair mutation that changed a protein).

It is common in those of African ancestry.

S will represent the normal dominant allele and s for the recessive sickle allele.

They are co‐dominant alleles.

SS is normal, Ss is not fatal, ss is debilitating, painful and often fatal.

Materials

Red beads

Blue beads

1 100mL Beaker

Procedure

1. Place 25 Red (S) and 25 Blue (s) beads into the 100 ml beaker and mix well.

2. Randomly (without looking) remove 2 beads. Repeat 10 times (without returning the beads

to the beaker), each time recording if it was a SS, Ss or ss.

3. Remove each ss from the population – they died.

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Lab 15: Population Genetics

4. The remaining beads survived and reproduced.

5. Count how many red and blue beads remained (separately) and place twice that number

back in the beaker.

6. Repeat the process 7 times.

Questions

1. What is the remaining ratio of alleles?

2. Have any been selected against?

3. Given enough generations, would you expect one of these alleles to completely disappear

from the population? Why or why not?

4. Would this be different if you started with a larger population? Smaller?

5. After hundreds or even thousands of generations both alleles are still common in those

of African Ancestry. How would you explain this?

6. The worldwide distribution of sickle gene matches very closely to the worldwide distribution

of Malaria.

(http://www.cdc.gov/malaria/distribution_epi/distribution.htm)

Is this significant? Why or why not?

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174


175


176


Introductory Biology

Appendix

Good Laboratory Techniques

177


178


Appendix: Good Lab Techniques

Good Laboratory Techniques

Science labs, whether at universities or in your home, are places of adventure and discovery. One of

the first things scientists learn is how exciting experiments can be. However, they must also realize

science can be dangerous without some instruction on good laboratory practices.

Read the protocol thoroughly before starting any new experiment.

You should be familiar with the action required

every step of the way.

Keep all work spaces free from clutter and dirty dishes.

Read the labels on all chemicals, and note the chemical

safety rating on each container. Read all MSDS (provided

on www.eScienceLabs.com).

Thoroughly rinse labware (test tubes, beakers, etc.) between

experiments. To do so, wash with a soap and hot

water solution using a bottle brush to scrub. Rinse completely

at least four times. Let air dry

Use a new pipet for each chemical dispensed.

Wipe up any chemical spills immediately. Check MSDSs

for special handling instructions (provided on

www.eScienceLabs.com).

A benchcoat will prevent any

spilled liquids from contaminating

the surface you work on.

A B C

Special measuring tools in make experimentation easier and more accurate in the lab.

A shows a beaker, B graduated cylinders, and C test tubes in a test tube rack.

179

.


Appendix: Good Lab Techniques



Use test tube caps or stoppers to cover test tubes

when shaking or mixing – not your finger!

When preparing a solution, refer to a protocol for

any specific instructions on preparation. Weigh out

the desired amount of chemicals, and transfer to a

beaker or graduated cylinder. Add LESS than the

required amount of water. Swirl or stir to dissolve

the chemical (you can also pour the solution back

and forth between two test tubes), and once dissolved,

transfer to a graduated cylinder and add the

required amount of liquid to achieve the final volume.

Disposable pipets aid in accurate

measuring of small volumes of

liquids. It is important to use a new

pipet for each chemical to avoid

contamination.

A molar solution is one in which one liter

(1L) of solution contains the number of grams equal to its molecular weight.

Ex:

1M = 11g CaCl x 110g CaCl/mol CaCl

(The formula weight of CaCl is 110g/mol)

A percent solution can be prepared by percentage of weight of chemical to 100ml

of solvent (w/v) , or volume of chemical in 100ml of solvent (v/v).

Ex:

20g NaCl + 80ml H 2 O = 20% w/v NaCl solution

Concentrated solutions, such as 10X, or ten times the normal strength, are diluted

such that the final concentration of the solution is 1X.

Ex:

To make a 100ml solution of 1X TBE from a 10X solution:

10ml 10X TBE + 90ml water = 100ml 1X TBE

Always read the MSDS before disposing of a chemical to insure it does not require

extra measures. (provided on www.eScienceLabs.com)

Don’t pour unused chemical back into the original bottle.

Avoid prolonged exposure of chemicals to direct sunlight and extreme temperatures.

180


Lab Appendix: 22: Plant Good Reproduction Lab Techniques

Immediately secure the lid of a chemical after use.

Prepare a dilution using the following equation:

Where c 1 is the concentration of the original solution, v 1 is the volume of the original solution,

and c 2 and v 2 are the corresponding concentration and volume of the final solution.

Since you know c 1 , c 2 , and v 2 , you solve or v 1 to figure out how much of the original solution

is needed to make a certain volume of a diluted concentration.


If you are ever required to smell a chemical, always waft a gas toward you, as shown in the

figure below.. This means to wave your hand over the chemical towards you. Never directly

smell a chemical. Never smell a gas that is toxic or otherwise dangerous.






Use only the chemicals needed for the activity.

Keep lids closed when a chemical is not being used.

When diluting an acid, always pour the acid into the water. Never pour water into an acid.

Never return excess chemical back to the original bottle. This can contaminate the chemical

supply.

Be careful not to interchange lids between different chemical bottles.

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Lab Appendix: 28: Ecological Good Lab Interactions Techniques




When pouring a chemical, always hold the lid of the chemical bottle between your fingers.

Never lay the lid down on a surface. This can contaminate the chemical supply.

When using knives or blades, always cut away from yourself.

Wash your hands after each experiment.

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1500 West Hampden Avenue

Building 5 Unit H

Sheridan, CO 80110

888.375.5487 • www.esciencelabs.com

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